Woody plants having improved growth properties

ABSTRACT

The invention relates to a method for producing a genetically modified plant or woody plant with improved growth properties (in terms of biomass and wood quality) as compared to a corresponding non-genetically modified wild type plant or woody plant, said method comprising altering the level of expression of a polypeptide in a plant cell or woody plant cell; a plant or woody plant; or a part thereof.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a divisional of U.S. patent application Ser. No.15/536,631, which entered the U.S. National Stage on Jun. 15, 2017, fromPCT/SE2015/051396, filed Dec. 29, 2015, which claims priority to DenmarkPatent Application No. PA201470833, filed Dec. 29, 2014, the disclosuresof which are incorporated herein by reference in their entirety.

SUBMISSION OF SEQUENCE LISTING AS ASCII TEXT FILE

The content of the following submission on ASCII text file isincorporated herein by reference in its entirety: a computer readableform (CRF) of the Sequence Listing (file name: 616562022710SeqList.txt,date recorded: Jan. 9, 2020, size: 661 KB).

FIELD OF THE INVENTION

The invention relates to a method for producing a genetically modifiedplant or woody plant with improved growth properties (in terms ofbiomass, wood quality) as compared to a corresponding non-geneticallymodified wild type plant or woody plant, said method comprising alteringthe level of expression of a polypeptide in a plant or woody plant cell,a plant or woody plant, or a part thereof.

BACKGROUND TO THE INVENTION

Perennial plants such as long-lived trees and woody plants have a lifestyle considerably different from annual plants, such as Arabidopsis, inthat perennial plants such as trees has an indeterminate growth, whereasplants such as Arabidopsis terminate growth when the plant flowers.

Perennial plants can also cycle between periods of active growth anddormancy. The lifecycle of long-lived trees and woody plants differssignificantly from annual crops, which often translocate carbon andnitrogen to seeds. Due to these differences between annual crops andperennial plants, such as trees, it has in many instances been foundthat a model system such as Populus tremula×tremuloides is a superiorsystem for reliably finding genes, which can be used for increasingbiomass production in woody plants.

Plant growth at apical meristems results in the development of sets ofprimary tissues and in the lengthening of the stem and roots. Inaddition to this primary growth, trees undergo secondary growth andproduce secondary tissue “wood” from the cambium. This secondary growthincreases the girth of stems and roots.

There are several factors such as different gene products that mightneed to be altered in order to enhance biomass production in trees.Growth in height, diameter, stem volume and wood density are importanttraits to follow for increased growth and biomass production. In view ofthe need to provide perennial plants capable of enhanced growth andbiomass in a range of different environmental conditions, as well aschanging environmental conditions, there is a continual need to provideplants with different genetic traits (comprising different sets ofactive genes) that adapt the plants for growth under these conditions.

In general, high yield plants can be made by crossing different lines,selecting plants with the best growing properties, where seeds fromthese plants can then be selected and new crosses can be performed. Inthis process, plants with better growth properties can be identified.One problem with trees and woody plants is that they need to be severalyears old before they produce flowers and can be used for traditionalcrossing. This can be overcome by using various molecular biologytechniques.

This invention describes how expression of a set of genes can be alteredto create transgenic woody plants, which have improved growthproperties, improved biomass and higher yield compared to thecorresponding non-genetically modified wild type woody plant.

SUMMARY OF THE INVENTION

The present invention provides a method for producing a geneticallymodified plant or woody plant having increased biomass and/or woodquality (wood density and/or wood biodegradability) compared to acorresponding non-genetically modified plant or woody plant of the samespecies, said method comprising:

-   -   (a) enhancing the level of expression of at least one        polypeptide having an amino acid sequence selected from among        SEQ ID NO.: 2, 28 and 38 or an ortholog thereof, and/or        -   reducing the expression of at least one polypeptide having            an amino acid sequence selected from among SEQ ID NO.: 58,            74, 88, 98, 106 and 128 or an ortholog or paralog thereof in            a woody plant, a woody plant cell or a part thereof;        -   (b) generating and/or selecting a woody plant, woody plant            cell or a part thereof with increased biomass and/or wood            density as compared to a corresponding non-genetically            modified woody plant; and        -   (c) growing the woody plant, the woody plant cell or the            part thereof under conditions which permit development of a            woody plant.

In one embodiment of the method, the at least one polypeptide isselected from among:

-   -   (a) a polypeptide having an amino acid sequence selected from        among SEQ ID NO: 2, 28, 38, 58, 74, 88, 98, 106 and 128;    -   (b) an ortholog polypeptide to the polypeptide having SEQ ID NO:        2, said ortholog polypeptide having at least 70% amino acid        sequence identity to a sequence selected from among SEQ ID NO:        2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24 and 26;    -   (c) an ortholog polypeptide to the polypeptide having SEQ ID NO:        28, said ortholog polypeptide having at least 70% amino acid        sequence identity to a sequence selected from among SEQ ID NO:        28, 30, 32, 34 and 36;    -   (d) an ortholog polypeptide to the polypeptide having SEQ ID NO:        38, said ortholog polypeptide having at least 70% amino acid        sequence identity to a sequence selected from among SEQ ID NO:        38, 40, 42, 44, 46, 48, 52, 54 and 56;    -   (e) an ortholog polypeptide to the polypeptide having SEQ ID NO:        58, said ortholog polypeptide having at least 70% amino acid        sequence identity to a sequence selected from among SEQ ID NO:        58, 60, 62, 64, 66, 68, 70 and 72;    -   (f) an ortholog polypeptide to the polypeptide having SEQ ID NO:        74, said ortholog polypeptide having at least 70% amino acid        sequence identity to a sequence selected from among SEQ ID NO:        74, 76, 78, 80, 82, 84 and 86;    -   (g) an ortholog polypeptide to the polypeptide having SEQ ID NO:        88, said ortholog polypeptide having at least 70% amino acid        sequence identity to a sequence selected from among SEQ ID NO:        88, 90, 92, 94 and 96;    -   (h) an ortholog polypeptide to the polypeptide having SEQ ID NO:        98, said ortholog polypeptide having at least 70% amino acid        sequence identity to a sequence selected from among SEQ ID NO:        98, 100, 102 and 104;    -   (i) an ortholog polypeptide to the polypeptide having SEQ ID NO:        106, said ortholog polypeptide having at least 70% amino acid        sequence identity to a sequence selected from among SEQ ID NO:        106, 108, 110, 112, 114, 116, 118, 120, 122, 124 and 126; and    -   (j) an ortholog polypeptide to the polypeptide having SEQ ID NO:        128, said ortholog polypeptide having at least 70% amino acid        sequence identity to a sequence selected from among SEQ ID NO:        128, 130, 132, 134, 136 and 138.

In one embodiment of the method, the genetically modified woody plant isa hardwood tree selected from the group consisting of acacia,eucalyptus, hornbeam, beech, mahogany, walnut, oak, ash, willow,hickory, birch, chestnut, poplar, alder, aspen, maple, sycamore, ginkgo,a palm tree and sweet gum.

In one alternative embodiment of the method, the genetically modifiedwoody plant of the method is a conifer selected from the groupconsisting of cypress, Douglas fir, fir, sequoia, hemlock, cedar,juniper, larch, pine, redwood, spruce and yew.

In one embodiment of the method, the genetically modified plant of themethod is a crop plant, for example sugarcane, pumpkin, maize (corn),wheat, rice, barley, rye, rape, forage grass, beet, cassava, soybeans,potatoes and cotton.

The invention provides a genetically modified woody plant, havingincreased biomass and/or wood quality (wood density and/or woodbiodegradability) as compared to a corresponding non-geneticallymodified woody plant of the same species that is produced by the methodof the invention.

The invention further provides a genetically modified woody plant havingincreased biomass and/or wood density as compared to a correspondingnon-genetically modified woody plant of the same species, said planthaving an enhanced level of expression of at least one polypeptidehaving an amino acid sequence selected from among SEQ ID NO.: 2, 28 and38 or an ortholog/paralog thereof, and/or

reducing the expression of at least one polypeptide having an amino acidsequence selected from among SEQ ID NO.: 58, 74, 88, 98, 106 and 128 oran ortholog/paralog.

In one embodiment, the genetically modified woody plant is a hardwoodtree selected from the group consisting of acacia, eucalyptus, hornbeam,beech, mahogany, walnut, oak, ash, willow, hickory, birch, chestnut,poplar, alder, aspen, maple, sycamore, ginkgo, a palm tree and sweetgum.

In an alternative embodiment, the genetically modified woody plant ofthe method is a conifer selected from the group consisting of cypress,douglas fir, fir, sequoia, hemlock, cedar, juniper, larch, pine,redwood, spruce and yew.

In one embodiment of the method, the genetically modified plant of themethod is a crop plant, for example sugarcane, pumpkin, maize (corn),wheat, rice, barley, rye, rape, forage grass, beet, cassava, soybeans,potatoes and cotton.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1: Glucose production rates of wood samples obtained fromtransgenic aspen expressing construct 35S022 and wild-type aspen, wherethe samples were prepared without [WP] or with [AP] an acidpre-treatment step.

FIG. 2: Carbohydrate composition of wood samples obtained fromtransgenic aspen expressing construct 35S022 and wild-type aspen.

DEFINITIONS

The term “improved growth properties” should be understood as primarygrowth, including a lengthening of the stem and roots, as well assecondary growth of a woody plant including production of secondarytissue, “wood”, from the cambium and an increase in the girth of stemsand roots. One way of monitoring growth is by measuring the height andthe diameter of the stem and optionally calculating the volume of thestem and comparing it with a wild type population or with parentalcontrol of the woody plants of interest. Improved growth produces aplant with increased biomass. Wood density is a positive measure of woodquality.

The term “improved wood quality” should, in one aspect be understood asincreased biodegradability of wood; in particular the saccharificationyield obtainable from wood derived from a woody plant of the invention.In particular, the susceptibility of cellulose in a wood derived from awoody plant to enzymatic cleavage and deconstruction of the polymericwood structure, as measurable by the yield of soluble sugars released oncleavage and deconstruction, is a measure of wood quality. Anotheraspect of wood quality is wood density which influences factors such asstrength of both fibrous products and solid wood products. Wood densityalso influences paper yield and properties. Wood density is a key factorfor kraft pulp production.

By “conditions which permit development of a tree” is meant that thenormal growth of the non-genetically modified woody plant, i.e. thewoody plant should be grown in the normal climate zone of the woodyplant. The temperature, day light and access to water and nutrientsshould be the norm for the growth region. An advantage with an improvedgrowth of the genetically modified woody plant is that the improvementmay also affect the survival of the genetically modified woody plants inan environment in which the non-genetically modified woody plants doesnot grow. This is very important from a commercial point of view.

By “biologically active variant” of a polypeptide is meant apolypeptide, protein or a stretch of amino acids, which have the sameactivity as the chosen polypeptide, but a different amino acid sequence,i.e. a biologically active variant of a polypeptide can perform the sameenzymatic reaction to create the same activity.

By “ortholog” or “orthologous polypeptide” is meant a polypeptideexpressed by evolutionarily related gene that has a similar nucleic acidsequence, where the polypeptide has similar functional properties.Orthologous genes are structurally related genes, from differentspecies, derived by a speciation event from an ancestral gene. Relatedto orthologs are paralogs. Paralogous genes are structurally relatedgenes within a single plant species most probably derived by aduplication of a gene. The word ortholog and paralog are usedinterchangeably in the entire text and the text may use the termortholog/paralog, where it is difficult to distinguish between orthologsand paralogs. Several different methods are known by those of skill inthe art for identifying and defining these functionally homologoussequences. An ortholog, a paralog or a homologous gene may be identifiedby one or more of the methods described below.

“Orthologous genes” from different organisms have highly conservedfunctions and can be used for identification of genes that could performthe invention in the same way as the genes presented here. Paralogousgenes, which have diverged through gene duplication, may encode proteinretaining similar functions. Orthologous genes are the product ofspeciation, the production of new species from a parental species,giving rise to two or more genes with common ancestry and with similarsequence and similar function. These genes, are termed orthologousgenes, often have an identical function within their host plants and areoften interchangeable between species without losing function.Identification of an “ortholog” gene may be done by identifyingpolypeptides in public databases using the software tool BLAST with oneof the polypeptides encoded by a gene. Subsequently additional softwareprograms are used to align and analyse ancestry. The sequence identitybetween two orthologous genes may be low. Implementation of suchidentification and analysis methods is illustrated in the introductionto the Examples.

The terms “substantially identical” or “sequence identity” may indicatea quantitative measure of the degree of homology between two amino acidsequences or two nucleic acids (DNA or RNA) of equal length. When thetwo sequences to be compared are not of equal length, they are alignedto give the best possible fit, by allowing the insertion of gaps or,alternatively, truncation at the ends of the polypeptide sequences ornucleotide sequences. The “sequence identity” may be presented aspercent number, such as at least 40, 50%, 55,%, 60%, 65%, 70%, 75%, 80%,81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%,95%, 96%, 97%, 98%, or at least 99% amino acid sequence identity of theentire length, when compared and aligned for maximum correspondence, asmeasured using a sequence comparison algorithm or by visual inspection.

In certain aspects, substantial identity exists over a region of nucleicacid sequences of at least about 50 nucleic acid residues, such as atleast about 100, 150, 200, 250, 300, 330, 360, 375, 400, 425, 450, 460,480, 500, 600, 700, 800 such as at least about 900 nucleotides or suchas at least about 1 kb, 2 kb, or such as at least about 3 kb.

In some aspects, the amino acid substantial identity exists over anpolypeptide sequences length of 10, 15, 20, 25, 30, 35, 40, 45, 50, 55,60, 65, 70, 75, 80, 85, 90, 95, 100, 110, 120, 130, 140, 150, 160, 170,180, 190, 200, 250, 300, 350, 400, 450, 500, 600, 700 amino acids in thepolypeptide with a “sequence identity” as defined above.

The sequence identity of the polypeptides of the invention can becalculated as (N_(ref)−N_(dif))100/N_(ref), wherein N_(dif) is the totalnumber of non-identical residues in the two sequences when aligned andwherein N_(ref) is the number of residues in one of the sequences. Thesequence identity between one or more sequence may also be based onglobal alignments using the clustalW software. In one embodiment of theinvention, alignment is performed with the sequence alignment methodClustalW with default parameters. The parameter set preferably used arefor pairwise alignment: Gap open penalty: 10; Gap Extension Penalty:0.1, for multiple alignment, Gap open penalty is 10 and Gap ExtensionPenalty is 0.2. Protein Weight matrix is set on Identity. BothResidue-specific and Hydrophobic Penalties are “ON”, Gap separationdistance is 4 and End Gap separation is “OFF”, No Use negative matrixand finally the Delay Divergent Cut-off is set to 30%.

Preferably, the numbers of substitutions, insertions, additions ordeletions of one or more amino acid residues in the polypeptide ascompared to its comparator polypeptide is limited, i.e. no more than 1,2, 3, 4, 5, 6, 7, 8, 9, or 10 substitutions, no more than 1, 2, 3, 4, 5,6, 7, 8, 9, or 10 insertions, no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or10 additions, and no more than 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10deletions. Preferably the substitutions are conservative amino acidsubstitutions: limited to exchanges within members of group 1: Glycine,Alanine, Valine, Leucine, Isoleucine; group 2: Serine, Cysteine,Selenocysteine, Threonine, Methionine; group 3: proline; group 4:Phenylalanine, Tyrosine, Tryptophan; Group 5: Aspartate, Glutamate,Asparagine, Glutamine.

The terms “hybridization” and “hybridize” are used broadly to designatethe association between complementary or partly complementary nucleicacid sequences. Under “stringent hybridization conditions”, nucleic acidbase pairing will occur only between nucleic acid fragments that have ahigh frequency of complementary base sequences. The length of thepolynucleotide fragment also affects the hybridization. An example of“stringent hybridization conditions” can be using a polynucleotidesequence of at least 15, 20, 25, 30, 35, 40, 45, 50, 100, or at least200 consecutive nucleotide residues, which hybridizes in 5× salinesodium citrate (SSC) at 40° C., followed by one or more washes in 2×SSC,0.2% sodium dodecyl sulphate (SDS) at 65° C. Lower temperature willreduce the stringency. More details about hybridization methods arefound in the art.

By “altering” is meant altering the level or the activity of a geneproduct. In this way “altering” is used for modifying, increasing,decreasing, reducing but not abolishing the levels or the activity of agene product within the plant. It can also refer to changing theexpression of the genes presented here; which can be used to modify thedesired properties.

Approaches to obtaining altered levels or activity of a gene product canbe done by using the nucleic acid construct as described for theidentification of plants having altered growth characteristics ascompared to the wild-type. Such plants may for instance be naturallyoccurring variants or plants that have been modified genetically toexhibit altered growth properties. For such purposes, the nucleic acidsequences according to the invention can be used as targets to identifygenetic variation that can be exploited as markers in a breedingprogram, e.g. as a probe in conventional hybridization assays or as aprimer for specific amplification of nucleic acid fragments.

The phrase “regulatory nucleic acid sequences” refers to regulatorybinding sites, promoters, poly-A signals and the similar.

By “reducing the amount or activity” of a polypeptide is meant that thetranscription and/or processing of mRNA might be reduced, whereby thesubsequent translation of the mRNA into a functional polypeptide mayresult in a lower amount of the polypeptide. The polypeptide can beprotein or an enzyme. When the amount of an enzyme is reduced theactivity might be reduced.

By “increasing the amount or activity” of a polypeptide is meant thatthe transcription of mRNA might be increased, the mRNA processing mightbe affected, resulting in an increase of the mRNA, whereby thetranslation of the mRNA into a functional polypeptide may result in ahigher amount of the polypeptide. The polypeptide can be a protein ormore specifically an enzyme. When the amount of an enzyme is increasedthe activity might be increased. Increasing the amount or activity of apolypeptide can also be achieved by introducing a nucleic acid sequencesinto a host cell, expressing said nucleic acid sequences and translatingit into a functional polypeptide. The functional polypeptide might notnormally be present or only normally expressed from the endogenous geneat a lower level, in such cases the amount or activity of thepolypeptide/enzyme is increased.

By “over-expressing” or “increased expression” is meant that a nucleicacid sequence after its introduction into a host cell is expressed at ahigher level than that normally expressed from the endogenous host geneencoding said polypeptide or protein.

DETAILED DESCRIPTION OF THE INVENTION

1. A Method for Increasing the Biomass Yield and/or Wood Density of aPlant or Woody Plant

The present invention provides methods for producing a geneticallymodified plant or woody plant having increased growth; whereby the woodyplant product yields increased biomass and/or increased wood density.The genetically modified (GM) plant or woody plant provided by theinvention, is characterised by an increased height, diameter, stemvolume, wood density, or any combination thereof, when compared to anon-genetically modified (non-GM) wild type population or to a parentalplant or woody plant used as control. Increased growth of a woody plantmay result from increased primary growth, including lengthening of thestem and roots, as well as increased secondary growth, includingproduction of secondary tissue “wood” from the cambium giving rise to anincrease in the girth of stems and roots.

It has surprisingly been found that genetic modification of a plant orwoody plant causing an altered expression level of one or morepolypeptide selected from among STT74, STT681, STT632, STT153, STT258,STT387, STT543, STT793, and STT795, wherein the amino acid of saidpolypeptide is SEQ ID NO: 2, 28, 38, 58, 74, 88, 98, 106 and 128respectively, or an ortholog or paralog thereof, and wherein the alteredexpression of said one or more polypeptide produces a plant having anincreased biomass and/or increased wood density and/or wood quality. Anortholog or paralog of the polypeptide is a polypeptide having at least40%, 45%, 50%, 55%, 60%, 65% 70%, 75%, 80%, 81%, 82%, 83%, 84%, 85%,86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, atleast 99% or 100% amino acid sequence identity any one of SEQ ID NO: 2,28, 38, 58, 74, 88, 98, 106 and 128; or a portion of any one of SEQ IDNO: 2, 28, 38, 58, 74, 88, 98, 106 and 128, as defined below in respectof each sequence.

It is known in the art that polypeptides encoded by orthologous genesretain their functional properties when transgenically expressed inheterologous plants or woody plants. For example, the expression ofgenes, derived from Arabidopsis thaliana, in tobacco and in treesconfers the same phenotypic properties on the transformed plant. Thus apolypeptide that is an ortholog to one of those described herein (e.gSTT74, STT681, STT632) is expected to function in the same way andimprove the growth properties when over-expressed in woody plants. Theexpression of polypeptides encoded by orthologous genes in a woodyplant, according to the present invention, has particular value since itmakes it possible to improve the growth properties of a woody plant ofhigh economic value, but where the native gene encoding the polypeptideto be expressed is not known. Similarly, reduced expression of apolypeptide that is an ortholog to one of those described herein (e.g.STT153, STT258, STT387, STT543, STT793, and STT795) encoded by anorthologous gene in a GM woody plant of the invention, by virtue of itsconserved functional properties, is expected to improve the growthproperties of the GM woody plants.

In a one embodiment, the invention provides a method for increasing thebiomass and/or wood density and/or wood quality of a plant or woodyplant; wherein the plant or woody plant is genetically modified in orderto increase the level of expression of one or more polypeptide, whereinthe amino acid sequence of the polypeptide has at least 70% sequenceidentity to a sequence selected from among SEQ ID NO: 2, 28, 38(corresponding to STT74, STT681 and STT632 respectively), or an orthologor paralog thereof as defined below in respect of each sequence.

In a further embodiment, the invention provides a method for increasingthe biomass yield and/or wood density and/or wood quality of a plant orwoody plant; wherein the plant or woody plant is genetically modified inorder to decrease the level of expression of one or more polypeptide,wherein the amino acid sequence of the polypeptide has at least 70%sequence identity to a sequence selected from among SEQ ID NO: 58, 74,88, 98, 106, 128 (corresponding to STT153, STT258, STT387, STT543,STT793 and STT795, respectively) or an ortholog or paralog as definedbelow in respect of each sequence.

1.1 Enhanced Expression of a Polypeptide (STT74) Having SEQ ID NO 2, oran Ortholog or Paralog Thereof, in a Plant or Woody Plant

In one embodiment, enhancing the expression of a polypeptide (STT74)having SEQ ID NO: 2 or an ortholog or paralog thereof, in a GM plant orwoody plant confers enhanced growth when compared to a non-GM plant orwoody plant used as control, as measured as the height, and diameter andvolume of the plant; as well as enhanced wood density (see example 1)and wood quality (see example 11). Functional properties assigned to theexpressed polypeptide are those of a vesicle-associated membraneprotein.

In one embodiment, the amino acid sequence of the polypeptide has atleast 70% sequence identity to SEQ ID NO: 2, and is selected from amongSEQ ID NO: 2 (corresponding to Populus trichocarpa polypeptide encodedby gene POPTR_0019s13890); SEQ ID NO: 4 (corresponding to Populustrichocarpa polypeptide encoded by gene Potri.013G147800.2); or SEQ IDNO:6 (corresponding to Populus trichocarpa polypeptide encoded by genePotri.019G116400).

Alternatively, the polypeptide has at least 70% sequence identity toamino acid residues 60 to 267 of SEQ ID NO 2, and is selected from amongSEQ ID NO: 8 (corresponding to Populus trichocarpa polypeptide encodedby gene Potri.001G408200.1); SEQ ID NO: 10 (corresponding to Populustrichocarpa polypeptide encoded by gene Potri.004G033500); SEQ ID NO: 12(corresponding to Populus trichocarpa polypeptide encoded by gene Potri.011G041900.1); SEQ ID NO: 14 (corresponding to Populus trichocarpapolypeptide encoded by gene Potri. 011G126200.2); SEQ ID NO:16(corresponding to Eucalyptus grandis polypeptide encoded by geneF01073); SEQ ID NO:18 (corresponding to Eucalyptus grandis polypeptideencoded by gene K00911); SEQ ID NO:20 (corresponding to Eucalyptusgrandis polypeptide encoded by gene D00750); SEQ ID NO: 22(corresponding to Arabidopsis thaliana polypeptide encoded by geneAT4G05060); SEQ ID NO: 24 (corresponding to Arabidopsis thalianapolypeptide encoded by gene AT4G21450); and SEQ ID NO: 26 (correspondingto Arabidopsis thaliana polypeptide encoded by gene AT5G54110).

In a preferred embodiment a polypeptide having at least 70% sequenceidentity to amino acid residues 60 to 267 of SEQ ID NO 2, for expressionin a GM plant or woody plant to enhance growth when compared to a non-GMplant or woody plant used as control, is characterised by comprising allpeptides listed in Table 1, wherein the amino acid sequence of peptideSTT74pep1 has at least 70% sequence identity to the corresponding regionin SEQ ID NO: 2, the amino acid sequence of peptide STT74pep2 has atleast 80% sequence identity to the corresponding region in SEQ ID NO: 2,and the amino acid sequence of peptide STT74pep3 has at least 90%sequence identity to the corresponding region in SEQ ID NO: 2.

TABLE 1 Peptides defining the STT74 polypeptide Amino acid position inSeq ID No.: 2 Length - No. First Last amino acids STT74pep1 72 267 195STT74pep2 84 153 70 STT74pep3 122 132 10

1.2 Enhanced Expression of a Polypeptide (STT681) Having SEQ ID NO 28,or an Ortholog or Paralog Thereof, in a Plant or Woody Plant

In one embodiment, enhancing expression of a polypeptide (STT681) havingSEQ ID NO: 28, or an ortholog or paralog thereof, in a GM plant or woodyplant confers enhanced growth when compared to a non-geneticallymodified (GM) woody plant used as control, as measured as the height,and diameter and volume of the plant; as well as enhanced wood density(see example 2). Functional properties assigned to the expressedpolypeptide are those of a GTPase activating protein.

In one embodiment, the amino acid sequence of the polypeptide has atleast 70% sequence identity to SEQ ID NO: 28, and is selected from amongSEQ ID NO: 28 (corresponding to Populus trichocarpa polypeptide encodedby gene POPTR_0001s38090); SEQ ID NO:30 (corresponding to Populustrichocarpa polypeptide encoded by gene Potri.011G098500); SEQ ID NO:32(corresponding to Eucalyptus grandis polypeptide encoded by geneEucgr.D00176); SEQ ID NO:34 (corresponding to Arabidopsis thalianapolypeptide encoded by gene AT4G21160); and SEQ ID NO:36 (correspondingto Arabidopsis thaliana polypeptide encoded by gene AT4G05330).

In a preferred embodiment a polypeptide having at least 70% sequenceidentity to SEQ ID NO: 28, for expression in a GM plant or woody plantto enhance growth when compared to a non-GM plant or woody plant used ascontrol, is characterised by comprising all of the peptides listed inTable 2, wherein the amino acid sequence of each of peptide STT681pep1,peptide STT681pep2, peptide STT681pep3 and peptide STT681pep4respectively has substantial sequence identity, or is identical, to thecorresponding region in SEQ ID NO: 28.

TABLE 2 Peptides defining the STT681polypeptide Amino acid position inSeq ID No.: 28 Length - No. First Last amino acids STT681pep1 29 39 11STT681pep2 47 76 30 STT681pep3 183 229 47 STT681pep4 239 256 18

1.3 Enhanced Expression of a Polypeptide (STT632) Having SEQ ID NO: 38,or an Ortholog or Para Log Thereof, in a Plant or Woody Plant

In one embodiment, enhancing expression of a polypeptide (STT632) havingSEQ ID NO: 38, or an ortholog or para log thereof, in a GM plant orwoody plant confers enhanced growth when compared to a non-GM plant orwoody plant used as control, as measured as the height, and diameter andvolume of the plant; as well as enhanced wood density (see example 3).The expressed polypeptide functions as a transcription factor, andbelongs to the so called WRKY family, characterized by a conservedregion with the amino acids WRKY. While not wishing to be bound bytheory, the functional properties assigned to WRKY family polypeptides,that contribute to the observed increase in woody plant growth anddensity, includes enhancing stress tolerance, eg. heat and salttolerance.

In one embodiment, the amino acid sequence of the polypeptide has atleast 70% sequence identity to a sequence selected from among SEQ ID NO:38 (corresponding to Populus trichocarpa polypeptide encoded by genePOPTR_0013s14960 gene); SEQ ID NO: 40 (corresponding to Populustrichocarpa polypeptide encoded by gene Potri. 013G153400.1); SEQ IDNO:42 (corresponding to Populus trichocarpa polypeptide encoded by genePotri.006G105300.1); SEQ ID NO: 44 (corresponding to Populus trichocarpapolypeptide encoded by gene Potri.016G128300.1); SEQ ID NO:46(corresponding to Populus trichocarpa polypeptide encoded by genePotri.019G123500.2); SEQ ID NO: 48 (corresponding to Eucalyptus grandispolypeptide encoded by gene Eucgr.B04010). SEQ ID NO: 50 (correspondingto Eucalyptus grandis polypeptide encoded by gene Eucgr.K02940); SEQ IDNO: 52 (corresponding to Arabidopsis thaliana polypeptide encoded bygene AtWRKY25 (AT2G30250)); SEQ ID NO: 54 (corresponding to Arabidopsisthaliana polypeptide encoded by gene AtWRKY33 (AT2G38470)); and SEQ IDNO: 56 (corresponding to Arabidopsis thaliana polypeptide encoded bygene AtWRKY26 (AT5G07100)).

In a preferred embodiment a polypeptide having at least 50% sequenceidentity to SEQ ID NO: 38, for expression in a GM plant or woody plantto enhance growth when compared to a non-GM plant or woody plant used ascontrol, is characterised by comprising all of the peptides listed inTable 3, wherein the amino acid sequence of each of peptide STT632pep1,peptide STT632pep2, peptide STT632pep3, peptide STT632pep4, peptideSTT632pep5 and peptide STT632pep6 respectively, has substantial sequenceidentity, or is identical, to the corresponding region in SEQ ID NO: 38.

TABLE 3 Peptides defining the ST632 polypeptide Amino acid position inSeq ID No.: 38 Length - No. First Last amino acids STT632pep1 69 79 11STT632pep2 119 133 15 STT632pep3 263 284 22 STT632pep4 274 280 7STT632pep5 267 280 14 STT632pep6 263 324 62 STT632pep7 258 315 58STT632pep8 430 488 59

1.4 Decreased Expression of a Polypeptide (STT153) Having SEQ ID NO: 58,or an Ortholog or Para Log Thereof, in a Plant or Woody Plant

In one embodiment, decreased expression of a polypeptide (STT153) havingSEQ ID NO: 58, or an ortholog or para log thereof, in a GM plant orwoody plant confers enhanced growth when compared to a non-GM plant orwoody plant used as control, as measured as the height, and diameter andvolume of the plant; as well as enhanced wood density (see example 4).Functional properties assigned to the expressed polypeptide are those ofa zinc finger protein.

In one embodiment, amino acid sequence of the polypeptide, whoseexpression is decreased, has at least 70% sequence identity to asequence selected from among SEQ ID NO: 58 (corresponding to Populustrichocarpa polypeptide encoded by gene POPTR_0018s01490 (v3.0 updatedto Potri.018G029900)); SEQ ID NO: 60 (corresponding to Populustrichocarpa polypeptide encoded by gene Potri.006G251300.1); SEQ ID NO:62 (corresponding to Populus trichocarpa polypeptide encoded by genePotri.001G172700.1); SEQ ID NO: 64 (corresponding to Eucalyptus grandispolypeptide encoded by gene Eucgr. F02548.1); SEQ ID NO:66(corresponding to Eucalyptus grandis polypeptide encoded by geneEucgr.C02807.1); SEQ ID NO:68 (corresponding to Eucalyptus grandispolypeptide encoded by gene Eucgr. C01779.1); SEQ ID NO: 70(corresponding to Arabidopsis thaliana polypeptide encoded by geneAT5G25490.1); and SEQ ID NO: 72 (corresponding to Arabidopsis thalianapolypeptide encoded by gene AT3G15680.1).

In a preferred embodiment a polypeptide having at least 65% sequenceidentity to SEQ ID NO: 58, whose expression is reduced in a GM plant orwoody plant to enhance growth when compared to a non-GM plant or woodyplant used as control, is characterised by comprising all of thepeptides listed in Table 4, wherein the amino acid sequence of peptideSTT153pep1, peptide STT153pep2, peptide STT153pep3, peptide STT153pep4,peptide STT153pep5, peptide STT153pep6, and peptide STT153pep7respectively, has substantial sequence identity, or is identical, to thecorresponding region in SEQ ID NO: 58.

TABLE 4 Peptides defining the STT153polypeptide Amino acid position inSeq ID No.: 58 Length - No. First Last amino acids STT153pep1 1 30 30STT153pep2 38 92 55 STT153pep3 47 62 16 STT153pep4 56 87 32 STT153pep565 82 18 STT153pep6 113 147 35 STT153pep7 119 136 18

1.5 Decreased Expression of a Polypeptide (STT258) Having SEQ ID NO: 74,or an Ortholog or Paralog Thereof, in a Plant or Woody Plant

In one embodiment, decreased expression of a polypeptide having SEQ IDNO: 74, or an ortholog or paralog thereof, in a GM plant or woody plantconfers enhanced growth when compared to a non-GM plant or woody plantused as control, as measured as the height, and diameter and volume ofthe plant; as well as enhanced wood density (see example 5). Functionalproperties assigned to the expressed polypeptide are those of acalmodulin binding protein.

In one embodiment, the amino acid sequence of the polypeptide, whoseexpression is decreased, has at least 70% sequence identity to asequence selected from among SEQ ID NO: 74 (corresponding to Populustrichocarpa gene POPTR_0013s13090, (or v3.0 updated toPotri.013G127200,)); SEQ ID NO: 76 (corresponding to Populus trichocarpapolypeptide encoded by gene Potri.019G095700); SEQ ID NO: 78(corresponding to Populus trichocarpa polypeptide encoded by genePotri.019G112400.1); SEQ ID NO: 80 (corresponding to Eucalyptus grandispolypeptide encoded by gene Eucgr.H00308.1); SEQ ID NO: 82(corresponding to Eucalyptus grandis polypeptide encoded by geneEucgr.L00007.2); SEQ ID NO: 84 (corresponding to Arabidopsis thalianapolypeptide encoded by gene AT3G59690.1); and SEQ ID NO: 86(corresponding to Arabidopsis thaliana polypeptide encoded by geneAT2G43680.3).

In a preferred embodiment a polypeptide having at least 60% sequenceidentity to SEQ ID NO: 74, whose expression is reduced in a GM plant orwoody plant to enhance growth when compared to a non-GM plant or woodyplant used as control, is characterised by comprising all of thepeptides listed in Table 5, wherein the amino acid sequence of each ofpeptide STT258pep1, peptide STT258pep2, peptide STT258pep3, peptideSTT258pep4, peptide STT258pep5, peptide STT258pep6, peptide STT258pep7,peptide STT258pep8, peptide STT258pep9, and peptide STT258pep10respectively, has substantial sequence identity, or is identical, to thecorresponding region in SEQ ID NO: 74.

TABLE 5 Peptides defining the STT258 polypeptide Amino acid position inSeq ID No.: 74 Length - No. First Last amino acids STT258pep1 1 15 15STT258pep2 1 26 26 STT258pep3 33 85 53 STT258pep4 107 128 22 STT258pep5130 224 85 STT258pep6 153 183 30 STT258pep7 249 295 47 STT258pep8 410460 51 STT258pep9 463 482 20 STT258pep10 507 517 11

1.6 Decreased Expression of a Polypeptide (STT387) Having SEQ ID NO: 88,or an Ortholog or Para Log Thereof, in a Plant or Woody Plant

In one embodiment, decreased expression of a polypeptide having SEQ IDNO: 88, or an ortholog or paralog thereof, in a GM plant or woody plantconfers enhanced growth when compared to a non-GM plant or woody plantused as control, as measured as the height, and diameter and volume ofthe plant; as well as enhanced wood density (see example 6). Thefunctional properties annotated to the expressed polypeptide are thoseof the enzyme shikimate dehydrogenase.

In one embodiment, the amino acid sequence of the polypeptide, whoseexpression is decreased, has at least 70% sequence identity to asequence selected from among SEQ ID NO: 88 (corresponding to Populustrichocarpa gene Potri.013G029900 gene); SEQ ID NO: 90 (corresponding toPopulus trichocarpa polypeptide encoded by gene Potri.005G043400.1); SEQID NO:92 (corresponding to Eucalyptus grandis polypeptide encoded bygene Eucgr.B01770.1); SEQ ID NO:94 (corresponding to Eucalyptus grandispolypeptide encoded by gene Eucgr.H01214.1); and SEQ ID NO:96(corresponding to Arabidopsis thaliana polypeptide encoded by geneAT3G06350.1).

In a preferred embodiment a polypeptide having at least 55% sequenceidentity to SEQ ID NO: 88, whose expression is reduced in a GM plant orwoody plant to enhance growth when compared to a non-GM plant or woodyplant used as control, is characterised by comprising all of thepeptides listed in Table 6, wherein the amino acid sequence of each ofpeptide STT387pep1, peptide STT387pep2, peptide STT387pep3, peptideSTT387pep4, peptide STT387pep5, peptide STT387pep6, peptide STT387pep7,peptide STT387pep8, and peptide STT387pep9 has substantial sequenceidentity, or is identical, to the corresponding region in SEQ ID NO: 88.

TABLE 6 Peptides defining the STT387 polypeptide Amino acid position inSeq ID No.: 88 Length - No. First Last amino acids STT387pep1 14 25 12STT387pep2 36 46 10 STT387pep3 71 84 14 STT387pep4 90 110 21 STT387pep5189 206 18 STT387pep6 237 258 22 STT387pep7 306 338 32 STT387pep8 363393 31 STT387pep9 458 472 15

1.7 Decreased Expression of a Polypeptide (STT543) Having SEQ ID NO: 98,or an Ortholog or Paralog Thereof, in a Plant or Woody Plant

In one embodiment, decreased expression of a polypeptide having SEQ IDNO: 98, or an ortholog or paralog thereof, in a GM plant or woody plantconfers enhanced growth when compared to a non-GM plant or woody plantused as control, as measured as the height, and diameter and volume ofthe plant; as well as enhanced wood density (see example 7). Functionalproperties assigned to the expressed polypeptide are those of a2-oxoglutarate- and Fe (II)-dependent oxygenase.

In one embodiment, the amino acid sequence of the polypeptide, whoseexpression is decreased, has at least 70% sequence identity to asequence selected from among SEQ ID NO: 98 (corresponding to Populustrichocarpa gene Potri.009G107600); SEQ ID NO:100 (corresponding toEucalyptus grandis polypeptide encoded by gene Eucgr.I01206.1); SEQ IDNO:102 (corresponding to Arabidopsis thaliana polypeptide encoded bygene AT3G19000.1); and SEQ ID NO:104 (corresponding to Arabidopsisthaliana polypeptide encoded by gene AT3G19010.1).

In a preferred embodiment a polypeptide having at least 65% sequenceidentity to SEQ ID NO: 98, whose expression is reduced in a GM plant orwoody plant to enhance growth when compared to a non-GM plant or woodyplant used as control, is characterised by comprising all of thepeptides listed in Table 7, wherein the amino acid sequence of each ofpeptide STT543pep1, peptide STT543pep2, peptide STT543pep3, peptideSTT543pep4, peptide STT543pep5, peptide STT543pep6, peptide STT543pep7,and peptide STT543pep8 has substantial identity, or is identical, to thecorresponding region in SEQ ID NO: 98.

TABLE 7 Peptides defining the STT543 polypeptide Amino acid position inSeq ID No.: 98 Length - No. First Last amino acids STT543pep1 47 65 19STT543pep2 85 94 10 STT543pep3 104 117 14 STT543pep4 163 183 21STT543pep5 191 292 102 STT543pep6 196 232 37 STT543pep7 238 257 20STT543pep8 261 292 32

1.8 Decreased Expression of a Polypeptide (STT793) Having SEQ ID NO:106, or an Ortholog or Paralog Thereof, in a Plant or Woody Plant

In one embodiment, decreased expression of a polypeptide having SEQ IDNO: 106, or an ortholog or paralog thereof, in a GM plant or woody plantconfers enhanced growth when compared to a non-GM plant or woody plantused as control, as measured as the height, and diameter and volume ofthe plant; as well as enhanced wood density (see example 8). Functionalproperties assigned to the expressed polypeptide are those of a smallGTP-binding protein, which is involved in cellular signal transduction.

In one embodiment, the amino acid sequence of the polypeptide, whoseexpression is decreased, has at least 70% sequence identity to SEQ ID NO8, and is selected from among SEQ ID NO: 106 (corresponding to Populustrichocarpa Potri.004G153400); SEQ ID NO: 108 (corresponding to Populustrichocarpa polypeptide encoded by gene Potri.009G115000.1); SEQ ID NO:110 (corresponding to Populus trichocarpa polypeptide encoded by genePotri.003G053400.1); SEQ ID NO: 112 (corresponding to Populustrichocarpa polypeptide encoded by gene Potri.001G182900.1); SEQ IDNO:114 (corresponding to Eucalyptus grandis polypeptide encoded by geneEucgr.G00442.1); SEQ ID NO:116 (corresponding to Eucalyptus grandispolypeptide encoded by gene Eucgr.F03029.1); SEQ ID NO:118(corresponding to a Eucalyptus grandis polypeptide encoded by geneEucgr.302962.1); SEQ ID NO:120 (corresponding to a Eucalyptus grandispolypeptide encoded by gene Eucgr.003821.1); SEQ ID NO:122(corresponding to an Arabidopsis thaliana polypeptide encoded by geneAT3G18820); SEQ ID NO:124 (corresponding to an Arabidopsis thalianapolypeptide encoded by gene AT1G49300.2); and SEQ ID NO:126(corresponding to an Arabidopsis thaliana polypeptide encoded by geneAT3G16100.1).

In a preferred embodiment a polypeptide having at least 70% sequenceidentity to SEQ ID NO: 106, whose expression is reduced in a GM plant orwoody plant to enhance growth when compared to a non-GM plant or woodyplant used as control, is characterised by comprising all of thepeptides listed in Table 8, wherein the amino acid sequence of each ofpeptide STT793pep1, peptide STT793pep2, peptide STT793pep3, peptideSTT793pep4, and peptide STT793pep5 has substantial sequence identity, oris identical, to the corresponding region in SEQ ID NO: 106.

TABLE 8 Peptides defining the STT793 polypeptide Amino acid position inSeq ID No.: 106 Length - No. First Last amino acids STT793pep1 4 29 26STT793pep2 32 49 18 STT793pep3 58 91 34 STT793pep4 110 142 33 STT793pep5145 161 17

1.9 Decreased Expression of a Polypeptide (STT795) Having SEQ ID NO:128, or an Ortholog or Paralog Thereof, in a Plant or Woody Plant

In one embodiment, decreased expression of a polypeptide having SEQ IDNO: 128, or an ortholog or paralog thereof, in a GM plant or woody plantconfers enhanced growth when compared to a non-GM plant or woody plantused as control, as measured as the height, and diameter and volume ofthe plant; as well as enhanced wood density (see example 9). Functionalproperties assigned to the expressed polypeptide are those of acalcium-binding protein with an EF-hand motif.

In one embodiment, the amino acid sequence of the polypeptide, whoseexpression is decreased, has at least 70% sequence identity to asequence selected from among SEQ ID NO: 128 (corresponding to Populustrichocarpa Potri.002G008600); SEQ ID NO: 130 (corresponding to Populustrichocarpa Potri.T102700.1); SEQ ID NO: 132 (corresponding to Populustrichocarpa Potri.005G253000.1); SEQ ID NO:134 (corresponding to aEucalyptus grandis polypeptide encoded by gene Eucgr.F01786.1); SEQ IDNO:136 (corresponding to an Arabidopsis thaliana polypeptide encoded bygene AT1G20760.1); and SEQ ID NO:138 (corresponding to an Arabidopsisthaliana polypeptide encoded by gene AT1G21630.1).

In a preferred embodiment a polypeptide having at least 55% sequenceidentity to SEQ ID NO: 128, whose expression is reduced in a GM plant orwoody plant to enhance growth when compared to a non-GM plant or woodyplant used as control, is characterised by comprising all of thepeptides listed in Table 9, wherein the amino acid sequence of each ofpeptide STT795pep1, peptide STT795pep2, peptide STT795pep3, peptideSTT795pep4, STT795pep5, STT795pep6, peptide STT795pep7, peptideSTT795pep8, peptide STT795pep9, and peptide STT795pep10 has substantialsequence identity, or is identical, to the corresponding region in SEQID NO: 128.

TABLE 9 Peptides defining the STT795 polypeptide Amino acid position inSeq ID No.: 128 Length - No. First Last amino acids STT795pep1 5 95 91STT795pep2 9 52 44 STT795pep3 56 106 51 STT795pep4 246 268 23 STT795pep5249 268 20 STT795pep6 512 547 36 STT795pep7 557 587 31 STT795pep8 645673 29 STT795pep9 769 790 22 STT795pep10 899 919 21

2.0 Methods for Genetically Modifying the Expression of a Polypeptide ina Woody Plant of the Invention 2.1 Genetic Constructs and Methods forEnhancing Expression of a Polypeptide in a Plant or Woody Plant of theInvention

A nucleic acid molecule having a nucleic acid sequence encoding apolypeptide whose expression is to be enhanced in a plant or woody plant(see 1.1-1.3), may be produced synthetically. The sequence of thenucleic acid molecule will comprise a coding sequence for the respectivepolypeptide; and whose nucleotide sequence is preferably optimised forexpression in the respective plant or woody plant. An example of asuitable nucleic acid molecule encoding a polypeptide for enhancedexpression in a plant or woody plant according to the invention isprovided in the sequence listing. The nucleic acid molecule, encoding apolypeptide for use in the invention, is operably linked (fused) tocis-regulatory regions comprising a promoter nucleic acid molecule andpreferably also a terminator nucleic acid molecule. The promoter may,for example, be a constitutive promoter (e.g. CaMV 35S promoter) or aplant promoter of the native plant gene encoding the polypeptide of theinvention, or a tissue specific promoter. The terminator nucleic acidmolecule may be a CaMV 35S terminator.

A nucleic acid molecule, encoding a polypeptide for use in theinvention, operably linked to cis-regulatory regions, is introduced intoa nucleic acid construct (vector) to ensure efficient cloning in E. colior Agrobacterium strains, and which make it possible to stably transformplants. Such vectors include various binary and co-integrated vectorsystems, which are suitable for T-DNA-mediated transformation. Thevector systems are generally characterized by having at least the virgenes, which are required for Agrobacterium-mediated transformation, andT-DNA border sequences

2.2 Genetic Constructs and Methods for Reducing Expression of aPolypeptide in a Plant or Woody Plant of the Invention

The following methods serve to illustrate alternative means fordown-regulating or silencing the functional activity of polypeptide(STT153, STT258, STT387, STT543, STT793 and STT795 or an ortholog orparalog thereof, as defined in 1.4-1.9) in a plant cell of a plant orwoody plant, where the polypeptide is encoded by a nucleic acid moleculein the genome of the plant cell.

Antisense Transgenes for Silencing Expression of a Polypeptide

Down-regulating or silencing expression of either a naturally occurringgene expressing a polypeptide according to the invention (STT153,STT258, STT387, STT543, STT793 and STT795) or an ortholog or paralogthereof (as defined in 1.4-1.9), in a host plant can be obtained bytransforming a transgene comprising a nucleic acid molecule (as definedin 1.5 to 1.8) encoding said polypeptide or a part thereof, or amolecule whose nucleic acid sequence is the anti-sense sequence of anucleic acid molecule encoding said polypeptide or a part thereof, intothe host plant.

RNAi Transgenes for Silencing Expression of a Polypeptide

Down-regulating or silencing expression of a naturally occurring geneencoding a polypeptide according to the invention (STT153, STT258,STT387, STT543, STT793 and STT795 or an ortholog or paralog thereof, asdefined in 1.5-1.8) in a host plant can be obtained by “RNAinterference” or “RNAi”: RNAi employs a double-stranded RNA molecule ora short hairpin RNA to change the expression of a nucleic acid sequencewith which they share substantial or total homology.

Suppression of the naturally occurring gene by RNA interference can beachieved using a transgene comprising a nucleic acid moleculefunctioning as a promoter that is operably linked to a nucleic acidmolecule comprising a sense and anti-sense element of a segment(fragment) of genomic DNA or cDNA of the naturally occurring gene(comprising a nucleic acid molecule as defined above section 1). Thesense and anti-sense DNA components can be directly linked or joined byan intron or artificial DNA segment that can form a loop when thetranscribed RNA hybridizes to form a hairpin structure.

It may be preferable that there is complete sequence identity in thesequence used for down-regulation of expression of a target sequence,and the target sequence, although total complementarity or similarity ofsequence is not essential. One or more nucleotides per 25 nucleotides ofa given nucleic acid molecule may differ from the corresponding sequencein the target gene. Thus, a sequence employed in a down-regulation ofgene expression in accordance with the present invention may be awild-type sequence (e.g. gene) selected from those available, or avariant of such a sequence, such as ortholog or paralog genes of thepresented genes.

It is important to note that there are a large number of fragments witha length of 20 nucleotides that will function in an RNA interferenceprocess to reduce the expression or activity of a target gene. As anexample, for the gene STT153, which is 468 nucleotides long, some 448different 20 nucleotide long fragments exists, and it is expected thatmost of these 20 nucleotide long fragments will reduce the expression oractivity of the target gene by the RNA interference process. From apractical point-of-view, the interfering RNA molecule must be doublestranded molecule, which can be achieved by cloning the fragment ofinterest head-to-head or tail-to-tail forming an inverted repeatedsequence. Furthermore, the cloned DNA fragment forming the interferingRNA should be at least 17, 18, 19, 20, 21, 22, 23, 24 or 25 nucleotideslong. The cloned DNA fragment forming the interfering RNA may also be50, 100, 150, 200, 250, 300, 350, 400, 450, 500, 550, 600 or 650 up tothe full length nucleotides long. The cloned DNA fragment forming theinterfering RNA may be longer than the translated part of the mRNA ofthe gene. The present invention shows that both shorter DNA fragmentsand longer DNA fragments function unexpectedly well in the RNAinterference process to reduce the expression or activity of a targetgene, see Table 11.

Artificial microRNA for Silencing Expression of a Polypeptide

In another example, an artificial microRNA is constructed were apromoter drives the expression of an RNA molecule mimicking the functionof a microRNA and the sequence setting the gene specificity isrecombinantly introduced. In a particular embodiment of the presentinvention the nucleic acid construct, or recombinant DNA construct,further comprises a strong constitutive promoter in front of atranscribed cassette consisting of part of the target gene followed by aplant functional intron followed by the same part of the target gene inreverse orientation. The transcribed cassette is followed by aterminator sequence. The preferred vector is of such type wherein one ofthe nucleotide sequence of the invention is inserted in inverted repeatorientation.

Induced Mutation and TILLING for Modifying Expression of a Polypeptide

The down-regulation or silencing of expression of a polypeptideaccording to the invention (STT153, STT258, STT387, STT543, STT793 andSTT795 or an ortholog or paralog thereof, as defined in 1.5-1.8) in aplant or woody plant cell can be obtained by means of mutations, such aspoint mutations, in the genes encoding the polypeptides. Mutations canbe introduced randomly into the genome of a plant cell, and thenmutagenized plant cells can be selected by specific methods such likeTILLING (Targeting Induced Local Lesions IN Genomes). Plants and plantcells, in which expression of a respective gene is down-regulated orsilenced as the result of a chemically induced mutation in their genome,are to be considered to be “genetically modified”, and since they do notcomprise a transgene introduced into their genome they are notconsidered to be recombinant plants or plant cells.

T-DNA Insertion in a Gene for Silencing Expression of a Polypeptide

Down-regulation or silencing of expression of a gene encoding apolypeptide according to the invention (STT153, STT258, STT387, STT543,STT793 and STT795 or an ortholog or paralog thereof, as defined in1.5-1.8), in a plant cell can also be obtained by T-DNA mutagenesis,whereby the T-DNA is used to randomly introduce mutations in the plantgenome followed by selecting plants comprising (non-silent) silencingmutations in the endogenous genes. The plant, or plant cell, in whicheither the endogenous gene is mutated can later be identified by PCR orother high throughput technologies using a series of PCR primer pairsspanning the respective gene.

Site Directed Mutagenesis for Modifying the Expression of a Polypeptide.

Modifying the expression of a gene encoding a polypeptide according tothe invention (STT153, STT258, STT387, STT543, STT793 and STT795 or anortholog or paralog thereof, as defined in 1.1-1.9), can be performed bymutating parts of the gene regulatory sequences using the site-directedmutagenesis method of site-directed nucleases. Three such differenttechnologies are Talen's, engineered Zinc finger nucleases andCrisper-cas. The basic mechanism is to modify the nuclease such that itis directed to a unique or near unique target DNA sequence in the targetgene, the technology is then introduced into the cell and the nucleasewill cut at or near the target sequence, the plants own DNA repairmechanism will then repair the cut DNA and in doing so a mutation willbe produced in some cases. Individual plants with the mutation will thenbe identified and the selected plants will be tested for the desiredeffect, e.g. increased biomass production.

2.3 Methods for Introducing Genetic Constructs into a Plant or WoodyPlant by Transformation

Transformation of Plant Cells

In accordance with the present invention, the method comprisestransforming regenerable cells of a plant with a nucleic acid constructor recombinant DNA construct (as described in 2.1 and 2.2) andregenerating a transgenic plant from said transformed cell. Productionof stable, fertile transgenic plants is now a routine method.

Various methods are known for transporting the construct into a cell tobe transformed. Agrobacterium-mediated transformation is widely used bythose skilled in the art to transform tree species, in particularhardwood species such as poplar and Eucalyptus. Other methods, such asmicroprojectile or particle bombardment, electroporation,microinjection, direct DNA uptake, liposome mediated DNA uptake, or thevortexing method may be used where Agrobacterium transformation isinefficient or ineffective, for example in some gymnosperm species.

A person of skill in the art will realise that a wide variety of hostcells may be employed as recipients for the DNA constructs and vectorsaccording to the invention. Non-limiting examples of host cells includecells in embryonic tissue, callus tissue type I, II, and III,hypocotyls, meristem, root tissue, tissues for expression in phloem,leaf discs, petioles and stem internodes. Once the DNA construct orvector is within the cell, integration into the endogenous genome canoccur.

Selection of Transformed Plant Cells and Regeneration of Plant or WoodyPlants

Following transformation, transgenic plants are preferably selectedusing a dominant selectable marker incorporated into the transformationvector. Typically, such a marker will confer antibiotic or herbicideresistance on the transformed plants and selection of transformants canbe accomplished by exposing the plants to appropriate concentrations ofthe antibiotic or herbicide. A selection marker using the D-form ofamino acids and based on the fact that plants can only tolerate theL-form offers a fast, efficient and environmentally friendly selectionsystem.

Subsequently, a plant may be regenerated, e.g. from single cells, callustissue or leaf discs, as is standard in the art. Almost any plant can beentirely regenerated from cells, tissues and organs of the plant. Aftertransformed plants are selected and they are grown to maturity and thoseplants showing altered growth properties phenotype are identified.

2.4 Methods for Detecting Modified Expression of a Gene Encoding aPolypeptide in a Plant or Woody Plant of the Invention

Real-time RT-PCR can be used to compare gene expression, i.e. the mRNAexpression, levels in a GM plant or woody plant with the correspondingnon-GM plant or woody plant. The amount of the polynucleotides disclosedherein can be determined by analysing using Northern blots, sequencing,RT-PCR or microarrays.

Western blots with immune detection or gel shift assays can be used tomeasure the expression levels or amounts of a polypeptide expressed in aGM woody plant of the invention. Antibodies raised to the respectivepolypeptide may be used for specific immune-detection of the expressedpolypeptide in tissue derived from a woody plant.

3.0 A Genetically Modified Plant or Woody Plant of the Invention

A GM plant or woody plant having increased growth; whereby the plant orwoody plant product yields increased biomass and/or increased wooddensity, is characterised by an altered expression level of one or morepolypeptide, wherein the polypeptide has an amino acid selected fromamong STT74, STT681, STT632, STT153, STT258, STT387, STT543, STT793, andSTT795 having SEQ ID NO: 1-18 respectively, or an ortholog or paralogthereof (as defined in section 1.0).

In one embodiment, the GM woody plant is a tree; for example a hardwoodplant selected from the group consisting of acacia, eucalyptus,hornbeam, beech, mahogany, walnut, oak, ash, willow, hickory, birch,chestnut, poplar, alder, maple, sycamore, ginkgo, a palm tree and sweetgum.

In another embodiment, the GM woody plant belongs to the familyMyrtaceae and the family Salicaceae. Hardwood plants from the Salicaceaefamily, such as willow, poplar and aspen including variants thereof, areof particular interest, as these two groups include fast-growing speciesof tree or woody shrub which are grown specifically to provide timberand bio-fuel. Eucalyptus species are also examples of such fast growingtrees.

In another embodiment, the GM woody plant is a conifer, for example aconifer selected from the group consisting of cypress, Douglas fir, fir,sequoia, hemlock, cedar, juniper, larch, pine, redwood, spruce and yew.In an alternative embodiment, the GM woody plant is a fruit bearingplant for example one selected from the group consisting of apple, plum,pear, banana, orange, kiwi, lemon, cherry, grapevine and FIG. In analternative embodiment, the GM woody plant is selected from the groupconsisting of cotton, bamboo and rubber plants.

In yet a further embodiment, the invention provides the use of the genesSTT74, STT681, STT632, STT153, STT258, STT387, STT543, STT793, andSTT795 as candidate genes in marker assisted breeding.

4.0 Methods for Measuring Enhanced Growth, Wood Density and BiomassYield in a Plant or Woody Plant

The increased growth might be measured by the height, diameter and stemvolume. The density can be calculated and might be used as measurementfor the quality of the wood. As illustrated in the examples below, theheight of a GM woody plant of the invention was increased between 6 and15% compared to non-GM trees; and the diameter of the GM woody plant wasincreased between 2% to 22% compared to non-GM trees. The increased stemvolume was increased from between 3% and 73% in GM trees compared tonon-GM trees. The increased wood density was increased from between 1%and 15% of the stem density compared to non-GM trees. A summary of theimproved growth properties is found in Table 13 in the examples below.To verify that no changes of wood quality had occurred in the modifiedtrees, wood from trees was analysed with FTIR. The data was evaluatedusing a multivariate analysis tool; and no significant differences werenoted. In summary, growth properties can be improved without any loss inwood quality.

EXAMPLES

Methodology:

cloning, transformation, establishment of the expression levels,identification of ortholog genes and calculation of growth propertiesare detailed below.

I Choice of Genes and Selection of Orthologs

Candidate genes for use in changing and/or modifying the phenotype of aplant with regard to growth were extracted from data derived from thegenome sequencing of Populus trichocarpa. The selected genes (Table 10)were compared to, and in some instances corrected based on the sequenceof homologous genes in Arabidopsis thaliana and other plant species.

TABLE 10 Summary of genes, the corresponding nucleotide and proteinsequences and given construct names used in the invention. Amino acidSEQ Nucleotide SEQ Gene ID No. ID No. Plasmid Construction STT74 SEQ IDNo. 2 SEQ ID No. 1 35s022 STT681 SEQ ID No. 28 SEQ ID No. 27 TFSTT052STT632 SEQ ID No. 38 SEQ ID No. 37 TF0137 STT153 SEQ ID No. 58 SEQ IDNo. 57 KR458 STT258 SEQ ID No. 74 SEQ ID No. 73 KR546 STT387 SEQ ID No.88 SEQ ID No. 87 KR675 STT543 SEQ ID No. 98 SEQ ID No. 97 KR831 STT793SEQ ID No. 106 SEQ ID No. 105 KR892 STT795 SEQ ID No. 128 SEQ ID No. 127KR894

A method to identify putative orthologs and paralogs genes is to analysethe relationships between genes and their related genes in the same anddifferent plants species. A commonly accepted and widely used method toachieve this is the construction of phylogenetic trees. The phylogenetictree will reveal groupings of related proteins (clades) and depending onthe algorithm used, it may also show evolutionary distances. Proteinsequences for construction of trees are often picked from publiclyavailable genomic resources, such as Phytozome and NCBI, using a BLASTsearch. Any given search will provide the user with a number of hitsordered by a score which is determined by sequence similarity overconserved regions of the protein sequences. To construct a robust treeit is common practice to include sequences from several related species.The number of hits varies greatly, depending on the query sequence andsearch parameters (settings). A score cut-off is determined individuallyfor each search, usually by looking for significant drops inscore/sequence similarity. It is important to also include genes thatare closely related but that are not orthologous to your gene. Allselected sequences are aligned using a multiple sequence alignmentsoftware such as ClustalW. The alignment can then be used to construct aphylogenetic tree using software for phylogenetic analysis such as MEGA.The phylogenetic tree will show a visual representation of the proteinrelationships of the corresponding genes. It can be expected thatorthologs and paralogs will form distinct groups and thus be identified.

As an example, the above method was used to identify ortholog genes of35s022 (STT74). For this example the databases searched were the Populustrichocarpa v3.0, Eucalyptus grandis v1, both parts of the Phytozomedatabase, and Arabidopsis thaliana TAIR10 database. Homologous geneswere selected from the above searches and further analysed. ClustalW wasused as the alignment algorithm, and phylogenetic trees were constructedusing MEGA and the neighbour joining method. From this analysis thefollowing genes were identified as paralog and/or ortholog genes:AT4G05060, with amino acid sequence SEQ ID NO: 22; AT4G21450, with aminoacid sequence SEQ ID NO: 24; AT5G54110, with amino acid sequence SEQ IDNO: 26; Eucgr.F01073, with amino acid sequence SEQ ID NO: 16;Eucgr.K00911, with amino acid sequence SEQ ID NO: 18; Eucgr.D00750, withamino acid sequence SEQ ID NO: 20; Potri.001G408200, with amino acidsequence SEQ ID NO: 8; Potri.004G033500, with amino acid sequence SEQ IDNO: 10; Potri.011G041900, with amino acid sequence SEQ ID NO: 12;Potri.011G126200, with amino acid sequence SEQ ID NO: 14;Potri.013G147800, with amino acid sequence SEQ ID NO: 4; andPotri.019G116400, with amino acid sequence SEQ ID NO: 6.

These genes were further analysed to identify the amino acid sequenceidentity levels of amino acid sub-sequences of the encoded polypeptidesthat could be used to define ortholog genes, compared to otherhomologous genes naturally occurring in plants and thereby created byevolution, based on amino acid identity. The regions selected for thiswere regions that showed a clear drop in identity level between genesthat were identified as ortholog genes in the phylogenetic analysiscompared to the genes that were identified as not being ortholog genes.The result of amino acid identity analysis showed that genes encoding apolypeptide comprising peptides which have higher than 75% sequenceidentity to amino acids 72-267 of SEQ ID NO 2 and higher than 80%identity to amino acids 84-153 of SEQ ID NO 2 and higher than 90%identity to amino acids 122-132 of SEQ ID NO 2 respectively, can beidentified as ortholog genes encoding polypeptides orthologous to thepolypeptide of SEQ ID NO 2. The result was then tested by identifyingmore orthologs of other species using the NCBI non-redundant proteinsequence database and the identity levels above. These identifiedortholog genes were then confirmed by adding them to the phylogeneticanalysis.

II Cloning of a Gene for Expression of the STT74 Polypeptide

The gene, with the nucleic acid sequence SEQ ID No: 10, corresponding toPopulus trichocarpa polypeptide encoded by gene POPTR_0019s13890, wascloned into an over-expression vector under the control of the CaMV 35Spromoter, giving construct 35s022. To produce cDNA template, total RNAwas isolated from stem, leaf and bark tissue sampled from hybrid aspen(Populus tremula×tremuloides) clone T89 plants and reverse transcribedto cDNA using Superscript III First Strand Synthesis System(Invitrogen). The gene STT74 was then amplified by PCR with genespecific forward and reverse primers using Phusion high fidelity DNApolymerase (Finnzymes). The amplified gene was subcloned into a Gatewayentry vector (pDONR201) using BP recombination cloning (Invitrogen),followed by further subcloning into the binary over-expression vectorpK2GW7 using Gateway LR recombination cloning (Invitrogen), where thegene was placed under the control of the CaMV 35S promoter. The clonedgene was verified using restriction digest of the final pK2GW7 vectorwith insert.

III Cloning of Genes for Expression of STT632 and STT681 Polypeptides

The cDNA was obtained as described above. The transcription factor geneswere amplified from cDNA and subcloned into a Gateway entry vector(pENTR/D-TOPO) by TOPO cloning (Invitrogen), followed by furthersubcloning of the genes into the binary over-expression vector pK2GW7using Gateway LR recombination cloning system (Invitrogen), where thegenes were placed under the control of the CaMV 35S promoter. Theplasmid construct TFSTT052 contains the gene STT681 with the nucleicacid sequence SEQ ID No: 27, which corresponds to the similar Populustrichocarpa gene, POPTR_0001s38090 (v3.0 updated to Potri.001G37200).

The plasmid construct TF0137 contains the gene STT632 with the nucleicacid sequence SEQ ID No: 37; which correspond to POPTR_0013s14960 (v3.0updated to Potri.013G153400) gene, in Populus trichocarpa. The clonedgenes were 5′ and 3′ end sequenced and verified using standardtechniques before subsequent subcloning into the pK2GW7 vector.

IV Cloning Gene Fragments for Preparing RNAi Constructs

A fragment of each of the selected genes listed in Table 11, encodingSTT153, STT258, STT387, STT543, STT793, and STT795, was identifiedlocated in a region of low homology to related genes in order toincrease RNAi specificity. Gene-specific primers were designed from ESTsequence data to amplify the gene fragments. EST library clones wereused as template for PCR amplification. The amplified gene fragment wassubcloned into a Gateway entry vector (pDONR201) using BP recombinationcloning (Invitrogen), followed by subsequent subcloning into the binaryRNA interference vector pK7GWIWG2(I) using Gateway LR recombinationcloning (Invitrogen) according to manufacturer's recommendations. Afinal RNAi construct can be schematically described: [CaMV 35Spromoter]-[gene fragment (antisense direction)]-[intron]-[gene fragment(sense direction)]-[35S terminator]. When transcribed the invertedrepeats separated by the intron will form a double stranded hairpinshaped RNA molecule. The constructs were verified using restrictionenzyme digest of the final pK7GWIWG2(I) vector with insert.

TABLE 11 Fragments used for RNAi constructs Sequence of cloned Length ofcloned Gene RNAi fragment RNAi fragment STT153 SEQ ID No. 139 515 STT258SEQ ID No. 140 313 STT387 SEQ ID No. 141 254 STT543 SEQ ID No. 142 258STT793 SEQ ID No. 143 274 STT795 SEQ ID No. 144 261

Further details of each RNAi construct are as follows:

IVi RNAi Construct (KR458) for Reducing Expression of STT153

Two copies of a 515 nucleotide long DNA fragment, SEQ ID No: 139, wasinserted, as an inverted repeat in plasmid construct KR458. Thisfragment originates from a hybrid aspen cDNA from the EST cloneUB11CPC10. The gene down-regulated by the RNAi construct KR458 in poplarcorresponds to the gene POPTR_0018s01490 (v3.0 updated toPotri.018G029900) encoding the polypeptide of SEQ ID NO: 58 in theclosely related Populus trichocarpa.

IVii RNAi Construct (KR546) for Reducing Expression of STT258

Two copies of a 313 nucleotide long DNA fragment, SEQ ID No: 140, wasinserted, as an inverted repeat in plasmid construct KR546. Thisfragment originates from a hybrid aspen cDNA from the EST clone G079P71.The gene down-regulated by the RNAi construct KR546 in poplarcorresponds to the gene POPTR_0013s13090, (or v3.0 updated toPotri.013G127200, encoding the polypeptide of in the closely relatedPopulus trichocarpa having SEQ ID No: 74.

IViii RNAi Construct (KR675) for Reducing Expression of STT387

Two copies of a 254 nucleotide long DNA fragment, SEQ ID No: 141, wasinserted, as an inverted repeat in plasmid construct KR675. Thisfragment originates from a hybrid aspen cDNA from the EST clone A044P01.The gene down-regulated by the RNAi construct KR675 in poplarcorresponds to the gene Potri.013G029900 gene, encoding the polypeptideof SEQ ID No.: 88 in the closely related Populus trichocarpa.

IViv RNAi Construct (KR831) for Reducing Expression of STT543

Two copies of a 254 nucleotide long DNA fragment, SEQ ID No: 142, wasinserted, as an inverted repeat in plasmid construct KR831. Thisfragment originates from a hybrid aspen cDNA from the EST clone F129P33.The gene down-regulated by the RNAi construct KR831 in poplarcorresponds to the gene Potri.009G107600 encoding the polypeptide of SEQID No: 98 in the closely related Populus trichocarpa.

IVv RNAi Construct (KR892) for Reducing Expression of STT793

Two copies of a 274 nucleotide long DNA fragment, SEQ ID No: 143, wasinserted, as an inverted repeat in plasmid construct KR892. Thisfragment originates from a hybrid aspen cDNA from the EST cloneUB30CPG09. The gene down-regulated by the RNAi construct KR892 in poplarcorresponds to the gene Potri.004G153400 encoding the polypeptide of SEQID No: 106, in the closely related Populus trichocarpa.

IVvi RNAi Construct (KR894) for Reducing Expression of STT795

Two copies of a 261 nucleotide long DNA fragment, SEQ ID No: 144, wasinserted, as an inverted repeat in plasmid construct KR894. Thisfragment originates from a hybrid aspen cDNA from the EST cloneUB30CPG09. The gene down-regulated by the RNAi construct KR894 in poplarcorresponds to the gene Potri.002G008600 encoding the polypeptide of SEQID No: 128, in the closely related Populus trichocarpa.

V Plant Transformation

DNA constructs were transformed into Agrobacterium and subsequently intoHybrid aspen, where Populus tremula×tremuloides clone T89, also called“poplar”, was transformed and regenerated. Approximately 3-8 independentlines were generated for each construct. One such group of transgenictrees produced using one construct is hereafter called a “constructiongroup”, e.g. different transgenic trees emanating from one construct.

Each transgenic line within each construction group derives from adifferent transformation event and has most probably the recombinant DNAinserted into a unique location in the plant genome. This makes thedifferent transgenic lines within one construction group partlydifferent. For example it is known that different transformation eventswill produce plants with different expression levels of the geneproduct. It is also known that different levels of expression of a genewill result in different levels of phenotypic effects.

VI Plant Growth

The transgenic poplar lines were grown together with their wild-typecontrol (wt) trees, in a greenhouse under a photoperiod of 18 h and atemperature of 22° C./15° C. (day/night). The plants were fertilizedweekly. The plants were grown for 8-9 weeks before harvest. During thistime their height and diameter was measured one to two times per week.In a growth group a number of wild-type trees (typically 35-45 trees)and a number of transgenic trees comprising several construction groups(typically 6-20 construction groups) were grown in parallel in thegreenhouse under the same conditions. All comparisons between thewild-type trees and construction groups are made within each cultivationgroup.

VII Growth Analyses

To identify construction groups showing a significant differencecompared to the wild type population, data from each construction groupwas subjected to a number of growth data analyses of growth/biomass andwood density measurements.

After 8 to 9 weeks growth in the greenhouse the trees were harvested andsampled. Two principal types of harvests were used; either a generalsetup designed for e.g. chemical analysis, wood morphology analysis,gene expression analysis, wood density analysis and metabolomicsanalysis, or a second setup designed for dry weight measurements ofbark, wood, leafs and roots.

Measurements of plant height and diameter were recorded one to two timesper week during the cultivation and before harvest of the plants. Finalheight and diameter measurements were subsequently used to identifyconstruction groups with altered growth characteristics.

The volume of the stem of each individual plant was approximated fromfinal height and final diameter measurements using the formula forvolume of a cone.

${{Stem}\mspace{14mu} {volume}\mspace{14mu} {approximation}\text{:}\mspace{11mu} V} = \frac{\pi*r^{2}*h}{3}$

where: V=Volume; h=height (Final height), r=radius (Final diameter/2)

Average final volumes of each construction group population andcorresponding wild type population were subsequently calculated.

VIII Replanting and Re-Growing

In order to verify data reproducibility and for further analysis, all ora subset of construction groups lines with growth characteristics ofextra interest were selected based on growth data from the firstcultivation in the greenhouse, replanted and regrown under the sameconditions as in the first greenhouse cultivation. All selectedtransgenic poplar lines were regrown in triplicates. Suffix denotingreplant round and transgenic line replicate were added to the names ofthe construction group transgenic lines in order to keep them unique.

IX Wood Density Analyses

Wood density is an important trait for increasing biomass production. Anincrease in wood density increases the energy content per cubic metrereduces the volume of a fixed amount of biomass and hence, e.g. thevolume required to transport a fixed amount of biomass. Correspondingly,more biomass can be transported per volume. Therefore increased densityis of interest, even if total biomass is not increased. Increaseddensity could also be of benefit coupled to pulp and paper production.

A 5 cm long stem segment, sampled between 36 and 41 cm from the soilfrom each harvested plant and stored in a freezer (−20° C.) afterharvest, was used for density measurements. Samples to be analysed werethawed followed by removal of bark and pith. The weight (w) was measuredusing a balance and the volume (V) was determined using the principle ofArchimedes, where wood samples were submerged (using a needle) into abeaker (placed on a balance) with water. The recorded increase in weightis equivalent to the weight of the water pushed aside by the woodsample. Since the density of water is 1 g/cm³ it is also equivalent tothe volume of the wood samples. The samples were then dried in ovenfor >48 h at 60° C.

The dry weights (dw) were measured and the density (d) was calculatedaccording to:

$d = \frac{dw}{V}$

Samples from each construction group were compared to wild type samplesfrom the same cultivation.

X Analysis of Expression Levels

Real-time RT-PCR was used to compare construct gene expression levels ofthe construction group with corresponding wild type group. Theexpression level of 26S proteasome regulatory subunit S2 was used as areference to which construct gene expression was normalized. Thecomparative CT method was used for calculation of relative constructgene expression level, where the ratio between construction andreference gene expression level is described by(1+E_(target))−CT_(target)/(1+E_(reference))−CT_(reference), whereE_(target) and E_(reference) are the efficiencies of construct andreference gene PCR amplification respectively and CT_(target) andCT_(reference) are the threshold cycles as calculated for construct andreference gene amplification respectively.

The mRNA expression levels of the up- or down-regulated gene in each ofthe transformed lines is summarized in Table 12.

TABLE 12 Summary of mRNA expression levels. Steady-state level of mRNAtranscript of corresponding regulated gene in Construct used intransformed lines as compared to wild Gene transformation type control(by RT-PCR) Over-expression constructs STT74 35s022 16.5 to 95 timeshigher STT681 TFSTT052 9.9 to 38 times higher STT632 TF0137 1.5 to 1.8times higher Down-regulated expression constructs STT153 KR458 42.3% to85.7% STT258 KR546  6.5% to 97.5% STT387 KR675 37.0% to 74.4% STT543KR831  7.0% to 66.2% STT793 KR892 18.2% to 94.1% STT795 KR894 33.6% to35.1%XI Results from Greenhouse Tests

The genes/constructs/construction groups were analysed as describedabove. Data from the transgenic trees transformed with the selectedgenes are presented in the examples below, with growth and wood propertycharacteristics. For some construction groups the wood density has beenmeasured and for some construction groups density predictions have beenmade based on FT-IR analysis (see table headers).

It is noted here, and applies to all the following data, that the ratiobetween the transgenic and wild type populations shows the averagedifference between those groups of plants. However, it does notgenerally show the full potential of modifying the expression of thegene. This is because the calculations are based on different transgenicevents.

For an easy overview, the improved growth properties of transgenicPopulus tremula×tremuloides clone T89 plant, transformed with theplasmid constructs in Table 10, causing enhanced or reduced expressionof the protein encoded by the respective gene, are summarised in Table13. The percentage values are the ratio between the analysed constructand the wild type tree from the examples below.

TABLE 13 Summary of improved growth properties of GM woody plants. Thepresented values are the average for all the tested lines per construct.Best performing lines are probably better. Construct used in Genetransformation Height Diameter Volume Density Over-expression constructsSTT74 35s022 107-124% 102-122% 113-173% 115% STT681 TFSTT052 106% 100%103% 115% STT632 TF0137 115% 101% 119% 106% Down-regulated expressionconstructs STT153 KR458 109% 110% 131% N.D. STT258 KR546 116% 108% 135% 96% STT387 KR675 108-111% 101-121% 108-159% 100-102% STT543 KR831 105%104% 114% 111% STT793 KR892 106% 108% 123% 110% STT795 KR894 107% 108%124% 101%

The growth results for modulation of the expression of each gene in awoody plant are presented separately as examples of the invention.

Example 1: STT74

The expression level of mRNA from the gene STT74 was analysed indifferent lines of the construct 35S022. The increased expression levelwas between 16.5 to 95 times higher than the wild type expression levelwhen analysed with RT-PCR as described above.

The 35s022 construct has been cultured three times and producedtransgenic trees with significantly improved height, diameter,stem-volume and density values.

In the first cultivation with height, diameter, stem volume and densityincreases of in average 15%, 22%, 73% and 15% respectively, compared towild type trees.

In the second cultivation with a height increase of in average 14%compared to wild type trees.

In the third cultivation with height and stem volume increases of inaverage 12% and 25% respectively, compared to wild type trees.

Transgenic line 35s022BIO-1B has a height and stem volume increase of10% and 25% respectively, compared to wild type trees.

Transgenic line 35s022BIO-2B has a height and stem volume increase of24% and 46% respectively, compared to wild type trees.

Height Diameter Volume Density (cm) (mm) (cm3) (g/cm3) 35s022 Average147.2 9.6 36.0 0.314 Statistics Ratio 1.12 1.05 1.25 T-test (p-value)0.000025 0.016 0.00021 Number > Upper limit 7 2 6 Number < Lower limit 00 0 Score (++) (+ I) (++) Max/Avg 35s022BIOmax/WTavg 1.33 1.15 1.66 1.0735s022BIO Average 130.2 9.2 29.2 Line 1A Max 147.0 10.1 39.3 Min 118.08.5 22.2 STD 11.3 0.6 6.5 Number 5 5 5 T89 Average 121.4 8.98 25.8 Max142.0 10.1 33.6 Min 107.0 7.85 17.7 STD 6.9 0.56 4.0 Number 33 33 33Upper limit 135.5 10.1 33.9 Lower limit 107.4 7.8 17.7 Statistics Ratio1.07 1.02 1.13 T-test (p-value) 0.02 0.44 0.11 Number > Upper limit 1 01 Number < Lower limit 0 0 0 Score (Normal) (Normal) (Normal) 35s022BIOAverage 133.4 9.6 32.4 Line 1B Max 152.0 10.3 37.9 Min 112.0 8.5 20.9STD 14.6 0.9 7.0 Number 5 5 5 T89 Average 121.4 8.98 25.8 Max 142.0 10.133.6 Min 107.0 7.85 17.7 STD 6.9 0.56 4.0 Number 33 33 33 Upper limit135.5 10.1 33.9 Lower limit 107.4 7.8 17.7 Statistics Ratio 1.10 1.071.25 T-test (p-value) 0.004 0.047 0.0038 Number > Upper limit 3 2 3Number < Lower limit 0 0 0 Score (++) (+ I) (++) 35s022BIO Average 150.39.8 37.7 Line 2B Max 162.0 10.1 42.8 Min 139.0 9.3 31.1 STD 11.5 0.4 6.0Number 3 3 3 T89 Average 121.4 8.98 25.8 Max 142.0 10.1 33.6 Min 107.07.85 17.7 STD 6.9 0.56 4.0 Number 33 33 33 Upper limit 135.5 10.1 33.9Lower limit 107.4 7.8 17.7 Statistics Ratio 1.24 1.09 1.46 T-test(p-value) 0.00000014 0.026 0.000033 Number > Upper limit 3 0 2 Number <Lower limit 0 0 0 Score (++) (Normal) (++)

Example 2: STT681

The expression level of mRNA from the gene STT681 was analysed indifferent lines of the construct TFSTT052. The increased expressionlevel was 9.9 to 38 times of the wild type expression level whenanalysed with RT-PCR as described above.

The TFSTT052 construct has produced transgenic trees with a significantdensity increase of 15% compared to wild type trees.

Height Diameter Volume Density (cm) (mm) (cm3) (g/cm3) TFSTT052 Average136.3 9.2 30.8 0.326 Max 147.0 10.4 41.6 0.342 Min 114.0 7.4 16.1 0.286STD 11.9 1.1 8.5 0.021 Number 6 6 6 6 T89 Average 129.0 9.21 29.8 0.284Max 151.0 11.4 51.0 0.361 Min 56.0 3.50 1.8 0.222 STD 14.0 1.32 8.40.030 Number 55 55 55 41 Upper limit 157.1 11.9 46.7 0.344 Lower limit101.0 6.6 13.0 0.224 Statistics Ratio 1.06 1.00 1.03 1.15 T-test(p-value) 0.22 0.96 0.79 0.0018 Number > Upper limit 0 0 0 0 Number <Lower limit 0 0 0 0 Score (Normal) (Normal) (Normal) (+ P) Max/AvgTFSTT052max/WTavg 1.14 1.13 1.40 1.20

Example 3: STT632

The expression level of mRNA from the gene STT632 was analysed indifferent lines of the construct TF0137. The increased expression levelwas 1.5 to 1.8 times of the wild type expression level when analysedwith RT-PCR as described above.

The TF0137 construct has produced transgenic trees with a significantheight increase of in average 15% compared to wild type trees.

Height Diameter Volume Density (cm) (mm) (cm3) (g/cm3) TF0137 Average160.4 8.6 32.6 0.284 Max 200.0 9.6 47.8 0.332 Min 121.0 7.9 20.8 0.262STD 37.1 0.8 13.7 0.028 Number 5 5 5 5 T89 Average 139.7 8.56 27.3 0.268Max 155.0 9.9 39.0 0.321 Min 122.0 7.40 18.6 0.238 STD 8.6 0.71 5.60.019 Number 40 39 36 42 Upper limit 157.1 10.0 38.7 0.306 Lower limit122.3 7.1 15.9 0.229 Statistics Ratio 1.15 1.01 1.19 1.06 T-test(p-value) 0.0032 0.85 0.12 0.081 Number > 2 0 2 1 Upper limit Number < 10 0 0 Lower limit Score (++) (Normal) (+ I) (Normal) Max/Avg TF0137max/1.43 1.12 1.75 1.24 WTavg

Example 4: STT153

The expression level of mRNA from the gene STT153 was analysed indifferent lines of the construct KR458. The reduced expression level was42.3% to 85.7% of the wild type expression level when analysed withRT-PCR as described above.

The KR458 construct has produced transgenic trees with significantlyimproved diameter and stem volume values, with diameter and stem volumeincreases of in average 10% and 31% respectively, compared to wild typetrees.

Height Diameter Volume Density (cm) (mm) (cm3) (g/cm3) KR458 Average149.2 10.7 44.9 Max 156.0 11.7 54.5 Min 142.0 9.2 34.2 STD 4.8 0.9 7.4Number 6 6 6 T89 Average 136.6 9.75 34.3 Max 165.0 11.2 47.0 Min 109.08.10 22.0 STD 12.5 0.64 6.0 Number 38 38 38 Upper limit 162.0 11.0 46.5Lower limit 111.2 8.5 22.1 Statistics Ratio 1.09 1.10 1.31 T-test(p-value) 0.021 0.0029 0.00034 Number > 0 2 2 Upper limit Number < 0 0 0Lower limit Score (Normal) (++) (++) Max/Avg KR458max/ 1.14 1.20 1.59WTavg

Example 5: STT258

The expression level of mRNA from the gene STT258 was analysed indifferent lines of the construct KR546. The reduced expression level was6.5% to 97.5% of the wild type expression level when analysed withRT-PCR as described above.

The KR546 construct has produced transgenic trees with significantlyimproved height and stem volume values, with height and stem volumeincreases of in average 16% and 35% respectively, compared to wild typetrees.

Height Diameter Volume Density (cm) (mm) (cm3) (g/cm3) KR546ReTransAverage 146.9 10.4 41.9 0.293 Max 160.0 11.8 56.5 0.336 Min 115.0 9.124.9 0.256 STD 14.4 0.8 8.9 0.029 Number 8 8 8 8 T89 Average 127.1 9.5631.0 0.304 Max 145.0 10.9 43.7 0.376 Min 104.0 6.55 11.7 0.232 STD 11.40.94 7.5 0.036 Number 32 32 32 28 Upper limit 150.3 11.5 46.2 0.378Lower limit 103.9 7.6 15.8 0.231 Statistics Ratio 1.16 1.08 1.35 0.96T-test (p-value) 0.00017 0.031 0.0011 0.42 Number > Upper limit 4 1 2 0Number < Lower limit 0 0 0 0 Score (++) (Normal) (++) (Normal) Max/AvgKR546ReTransmax/WTavg 1.26 1.23 1.82 1.11

Example 6: STT387

The expression level of mRNA from the gene STT387 was analysed indifferent lines of the construct KR675. The reduced expression level was37.0% to 74.4% of the wild type expression level when analysed withRT-PCR as described above.

The KR675 construct has been cultured two times and produced transgenictrees with significantly improved height, diameter and stem volumevalues.

In the first cultivation with height, diameter and stem volume increasesof in average 11%, 21% and 59% respectively, compared to wild type.

In the second cultivation with a height increase of 8% compared to wildtype trees.

Height Diameter Volume Density (cm) (mm) (cm3) (g/cm3) KR675 Average134.7 10.1 35.9 0.283 Max 150.0 10.5 42.5 0.308 Min 125.0 9.5 29.5 0.260STD 10.1 0.5 5.2 0.022 Number 6 6 6 6 T89 Average 121.1 8.33 22.6 0.283Max 138.0 10.4 36.7 0.339 Min 103.0 5.65 8.9 0.227 STD 9.3 1.10 6.70.031 Number 32 32 32 26 Upper limit 140.1 10.6 36.4 0.347 Lower limit102.0 6.1 8.9 0.219 Statistics Ratio 1.11 1.21 1.59 1.00 T-test(p-value) 0.0026 0.0006 0.000057 0.97 Number > Upper limit 2 0 3 0Number < Lower limit 0 0 0 0 Score (++) (+ P) (++) (Normal) Max/AvgKR675max/WTavg 1.24 1.25 1.88 1.09 KR675rp1 Average 150.5 8.6 29.4 0.269Max 161.0 9.2 34.1 0.289 Min 139.0 7.8 21.9 0.251 STD 8.1 0.4 3.8 0.011Number 11 8 8 11 T89 Average 139.7 8.56 27.3 0.268 Max 155.0 9.9 39.00.321 Min 122.0 7.40 18.6 0.238 STD 8.6 0.71 5.6 0.019 Number 40 39 3642 Upper limit 157.1 10.0 38.7 0.306 Lower limit 122.3 7.1 15.9 0.229Statistics Ratio 1.08 1.01 1.08 1.00 T-test (p-value) 0.00055 0.81 0.320.87 Number > Upper limit 3 0 0 0 Number < Lower limit 0 0 0 0 Score(++) (Normal) (Normal) (Normal) Max/Avg KR675rp1max/WTavg 1.15 1.07 1.251.08 KR675rp1 Average 153.7 8.6 31.2 0.273 Line 3A Max 161.0 8.7 31.50.280 Min 141.0 8.6 30.8 0.270 STD 11.0 0.0 0.5 0.006 Number 3 2 2 3 T89Average 139.7 8.56 27.3 0.268 Max 155.0 9.9 39.0 0.321 Min 122.0 7.4018.6 0.238 STD 8.6 0.71 5.6 0.019 Number 40 39 36 42 Upper limit 157.110.0 38.7 0.306 Lower limit 122.3 7.1 15.9 0.229 Statistics Ratio 1.101.01 1.14 1.02 T-test (p-value) 0.011 0.9 0.35 0.61 Number > Upper limit2 0 0 0 Number < Lower limit 0 0 0 0 Score (+ I) (Normal) (Normal)(Normal)

Example 7: STT543

The expression level of mRNA from the gene STT543 was analysed indifferent lines of the construct KR831. The reduced expression level was7.0% to 66.2% of the wild type expression level when analysed withRT-PCR as described above.

The KR831 construct has produced transgenic trees with a significantdensity increase of in average 11% compared to wild type trees.

Height Diameter Volume Density (cm) (mm) (cm3) (g/cm3) KR831 Average117.4 7.7 19.1 0.297 Max 130.0 9.2 26.6 0.355 Min 100.0 6.2 9.9 0.265STD 10.2 1.2 6.8 0.036 Number 7 7 7 7 T89 Average 112.0 7.47 16.7 0.266Max 125.0 8.7 23.4 0.358 Min 93.0 5.40 7.5 0.219 STD 8.3 0.77 4.1 0.022Number 32 32 32 30 Upper limit 129.0 9.0 25.1 0.312 Lower limit 95.0 5.98.3 0.221 Statistics Ratio 1.05 1.04 1.14 1.11 T-test (p-value) 0.140.47 0.23 0.0075 Number > 1 1 2 3 Upper limit Number < 0 0 0 0 Lowerlimit Score (Normal) (Normal) (+ I) (++) Max/Avg KR831max/ 1.16 1.231.59 1.33 WTavg

Example 8: STT793

The expression level of mRNA from the gene STT793 was analysed indifferent lines of the construct KR892. The reduced expression level was18.2% to 94.1% of the wild type expression level when analysed withRT-PCR as described above.

The KR892 construct has produced transgenic lines with height and stemvolume increases of up to 17% and 55% respectively, compared to the wildtype population average.

Density (g/cm3) Height Diameter Volume Prediction (cm) (mm) (cm3) fromFT-IR KR892 Average 158.3 9.1 34.6 0.315 Max 175.0 9.9 43.6 0.362 Min147.0 7.4 20.8 0.254 STD 12.2 0.9 7.8 0.036 Number 7 7 7 7 T89 Average150.0 8.4 28.2 0.286 Max 173.0 9.6 47.7 0.328 Min 130.0 7.3 18.8 0.240STD 10.6 0.7 6.0 0.025 Number 21 21 21 22 Upper limit 172.1 9.9 40.70.337 Lower limit 127.8 7.0 15.6 0.235 Statistics Ratio 1.06 1.08 1.231.10 T-test (p-value) 0.09386 0.05448 0.03248 0.02320 Number > Upperlimit 2 1 2 1 Number < Lower limit 0 0 0 0 Score (+ I) (Normal) (+ I)(Normal) Max/Avg KR892max/WTavg 1.17 1.18 1.55 1.27

Example 9: STT795

The expression level of mRNA from the gene STT795 was analysed indifferent lines of the construct KR894. The reduced expression level was33.6% to 35.1% of the wild type expression level when analysed withRT-PCR as described above.

The KR894 construct has produced transgenic trees with significantlyimproved height, diameter and stem volume values, with height, diameterand stem volume increases of in average 7%, 8% and 24% respectively,compared to wild type trees.

Density (g/cm3) Height Diameter Volume Prediction (cm) (mm) (cm3) fromFT-IR KR894 Average 158.2 9.3 35.8 0.292 Max 164.0 9.8 40.8 0.312 Min150.0 8.8 32.4 0.267 STD 5.2 0.4 3.5 0.018 Number 5 5 5 5 T89 Average148.4 8.6 28.8 0.291 Max 159.0 9.6 38.4 0.325 Min 132.0 7.7 21.1 0.250STD 7.5 0.5 4.0 0.019 Number 31 31 31 31 Upper limit 163.8 9.6 36.90.329 Lower limit 133.1 7.6 20.6 0.252 Statistics Ratio 1.07 1.08 1.241.01 T-test (p-value) 0.00867 0.00377 0.00076 0.84974 Number > Upperlimit 1 2 2 0 Number < Lower limit 0 0 0 0 Score (+ P) (++ ) (+ +)(Normal) Max/Avg KR894max/WTavg 1.10 1.14 1.42 1.07

Example 10: Field Trial of Hybrid Aspen with Lines Comprising aTransgene Having SEQ ID NO: 1 and Encoding a STT74 Polypeptide

Hybrid aspen field trials were established to further study the improvedgrowth properties of the transgenic trees under field conditions. Eachfield trial contains plants from 7 to 16 gene constructs and about 20%wild type (wt) reference plants. For each gene construct three to sixtransgenic plant lines, each derived from different transformationalevents, were selected for field trial. The transgenic plant lines weremultiplied in 8 to 20 replicates each. The transgenic plant lines weredistributed in field following a randomized block design. In the fieldall plants were separated in a 3×3 meter coordinate system to make asingle cell plant design. Whenever possible the field trials weredivided into two separate experimental sites, which were distant fromeach other and differ somewhat in environmental characteristics. Thefield sites were prepared and homogenized according to standardagricultural procedures such as disc harrowing and glyphosate basedherbicide treatment. The hybrid aspen field trials started 2011 and isplanned to proceed for 5 years. Within this time growth propertiesshould be regularly monitored and analysed.

After two growth seasons the preliminary results show an increasedheight of up to 29% between transgenic plant lines and wildtype, seealso Table 14.

TABLE 14 Increased growth of hybrid aspen after two growth seasons.Plant T-test Dunnett's Lines Mean Ratio LSqMean Ratio P-value P-ValueHeight August 2012 35s022F3-1A 92.0 0.97 90.6 0.95 0.7564 1.000035s022F3-1B 111.3 1.18 111.3 1.17 0.0346 0.4781 35s022F3-2A 113.5 1.20113.5 1.20 0.0176 0.1956 35s022F3-2B 101.0 1.07 101.0 1.06 0.4207 1.0000T89-wt 94.7 1.00 94.9 1.00 * * Height September 2013 35s022F3-1A 157.60.92 156.5 0.91 0.4227 1.0000 35s022F3-1B 220.1 1.29 220.1 1.29 0.00220.0397 35s022F3-2A 196.9 1.15 196.9 1.15 0.1156 0.9942 35s022F3-2B 180.11.05 180.1 1.05 0.5777 1.0000 T89 171.1 1.00 171.7 1.00 * *

Example 11: Properties of the Wood of a GM Wood Plant ExpressingConstruct 355022 Encoding a STT74 Polypeptide

Wood samples from three lines of GM aspen lines transformed with the35s022 construct, expressing a polypeptide (STT74) having SEQ ID NO: 1,were analysed to determine their susceptibility to pre-treatment andenzymatic saccharification. The transgenic lines were referred to as 1A,1B, and 2B or, alternatively, H12.1, H12.2, and H12.3. As a control, T89hybrid aspens referred to as “wild-type” were used. Pre-treatment wasperformed using acid hydrolysis, a state-of-the-art method for woodybiomass.

Experimental Protocol

Pre-treatment:

Wood of wild-type and transgenic aspen lines (T89 and transgenic lines1A, 1B, and 2B) was milled to a powder. Fifty mg of wood powder in areaction mixture with a total weight of 1000 mg were pre-treated using asingle-mode microwave system (Initiator Exp, Biotage, Uppsala, Sweden)using an acid catalyst [1% (w/w) sulphuric acid]. The pre-treatment wasperformed for 10 min at 165° C. The solid and liquid fractions wereseparated by centrifugation for 15 min at 14,100 g in pre-weighedmicro-centrifuge tubes. The liquid fraction, referred to as thepre-treatment liquid, was collected for analysis, while the solidfraction was washed twice with one ml of deionized water and once withone ml of sodium citrate buffer (50 mM, pH 5.2) prior to enzymatichydrolysis. The weight of the residual washed solids from thepre-treatment was determined.

Enzymatic Hydrolysis:

Sodium citrate buffer (50 mM, pH 5.2) and 50 mg of an enzyme cocktailconsisting of equal proportions of Celluclast 1.5 L and Novozyme 188[obtained from Sigma-Aldrich (St. Louis, Mo., USA)] were added topre-treated or non-pre-treated wood so that the total weight of thereaction mixture was 1000 mg. Reaction mixtures with wood that had notbeen pre-treated consisted of 50 mg of milled wood, 900 mg of the sodiumcitrate buffer, and 50 mg of the enzyme cocktail. The reaction mixtureswere incubated for 72 h at 45° C. in an orbital shaker (Ecotronincubator shaker, Infors,

Bottmingen, Switzerland) set at 170 rpm. Samples for analysis of glucoseformation during the early phase of the reaction (the glucose productionrate, GPR) were taken after 2 h. The liquid remaining after 72 h wasanalysed using high-performance anion-exchange chromatography (HPAEC).

Analysis of Hydrolysates:

The glucose concentrations during the early phase of the enzymaticreaction (the first 2 h) were measured using a glucometer. The yields ofmonosaccharide sugars (arabinose, galactose, glucose, xylose andmannose) in the pretreatment liquid and in the samples taken after 72 hof enzymatic hydrolysis were determined by using HPAEC. The HPAEC system(Ion Chromatography System ICS-3000, Dionex, Sunnyvale, Calif., USA) wasequipped with a PAD (pulsed amperometric detection) unit. The separationwas performed using CarboPac PA20 column (3×150 mm) (Dionex) equippedwith a CarboPac PA20 guard column (3×30 mm) (Dionex). Prior toinjection, the samples were filtered through 0.2 μm nylon filters. Avolume of 10 μl was loaded. Elution of sugars was performed with a 2 mMsolution of sodium hydroxide during 27 min, followed by regenerationwith 100 mM sodium hydroxide for 5 min, and equilibration with 2 mMsodium hydroxide for 15 min. The flow rate was 0.4 ml min-1. Pulsedamperometric detection of monosaccharides was performed with thedetector set on Gold Standard PAD waveform and with Ag/AgCl as referenceelectrode. Peaks were identified and quantified by comparison ofstandards containing arabinose, galactose, glucose, xylose, and mannose(Sigma-Aldrich). The sugar yields in the pre-treatment liquid and in theenzymatic hydrolysates are reported as g of sugar per g of wood afterpre-treatment and after 72 h of enzymatic hydrolysis, respectively.

Acetic Acid Analysis:

The concentrations of acetic acid (acetic acid in the pre-treatmentliquid, and acetic acid in enzymatic hydrolysate) were determined byusing the ICS-3000 system and the conductivity detector (Dionex).Separation was performed with an AS15 (4×250 mm) separation columnequipped with an AG15 (4×50 mm) guard column (Dionex). The mobile phaseconsisted of a 35 mM solution of sodium hydroxide (Sodium HydroxideSolution for IC, Sigma-Aldrich), and the flow rate was 1.2 ml min-1.

Carbohydrate Analysis:

One hundred mg (dry weight) of the wood powder were hydrolysed withsulphuric acid [3 ml, 72% (w/w)] for 1 h at 30° C. The reaction mixturewas diluted to 2.5% sulphuric acid using deionized water and wasautoclaved for 1 h at 120° C. After centrifugation (14,000 g for 20min), the supernatant was collected and analysed with respect tomonosaccharide content using the ICS-3000 system.

Results

Glucose Production Rates:

The glucose production rates (GPR), i.e. the glucose formed during theinitial phase of the enzymatic reaction, is shown in FIG. 1. Withoutpre-treatment, the average GPR of the transgenic lines, 3.23 g L-1 h-1,was 13% higher than the GPR of the wild-type (2.87 g L-1 h-1), but thedifference was not statistically significant (P<0.05). Withpre-treatment, the average GPR of the three transgenic lines was 8.65 gL⁻¹ h⁻¹, while the GPR of the wild-type was only 7.45 g L-1 h-1. This16% increase in GPR of the transgenic lines was significantly (P<0.05)higher than the GPR of the wild-type.

Yields of Monosaccharides and Acetic Acid:

Table 1 shows the yields of monosaccharides and acetic acid in enzymatichydrolysates and in pre-treatment liquid. The table also shows themonosaccharide yields when sugars in different fractions are addedtogether, i.e. separately, in total, and divided into pentoses(arabinose and xylose) and hexoses (galactose, glucose and mannose).

Without pre-treatment, H12.1 showed 43% higher glucose yield and 28%higher mannose yield than the wild-type (P<0.05). The line H12.3 showed23% higher mannose yield than the wild-type (P<0.05). The averageglucose yield of the transgenic lines was 28% higher than that of thewild-type (P<0.06).

The differences in yield after pre-treatment were not significant (Table15). This can be attributed to the fact that less carbohydrate ishydrolysed in measurements of the GPR (both for non-pre-treated andpre-treated samples) and in measurements of the yield of non-pre-treatedsamples than in measurements of the yield of pre-treated samples.

TABLE 15 Yields of sugar and acids in enzymatic hydrolysates (after 72 hreaction) and in pretreatment liquid. Yield of Sugar/ P (3 Acetic Acidlines vs Saccharification (g g⁻¹) #H-H12.1 [5] #H-H12.2 [5] #H-H12.3 [3]T89 T 89) Without pretreatment Y_(Ara/w) 0.0042 ± 0.00(126)   0.0042 ±0.00(98.2)  0.0033 ± 0.00(80.5) 0.0028 ± 0.00(100) 0.024 Y_(Gal/w)0.0176 ± 0.00 (155)  0.0175 ± 0.00(155) 0.0145 ± 0.00(129) 0.0113 ±0.00(100) 0.16 Y_(Glu/w)  0.1962 ± 0.02(143)* 0.1670 ± 0.06(122) 0.1638± 0.02(120) 0.1365 ± 0.01(100) 0.053 Y_(Xyl/w) 0.0311 ± 0.00(112)  0.0271 ± 0.00(97.8) 0.0317 ± 0.00(114) 0.0277 ± 0.00(100) 0.524Y_(Man/w)  0.0091 ± 0.00(128)* 0.0073 ± 0.00(103)  0.0087 ± 0.00(123)*0.0071 ± 0.00(100) 0.133 Y_(acetic acid) 0.017 ± 0.002(101   0.017 ±0.002(102) 0.017 ± 0.001(101) 0.017 ± 0.002(100) 0.773 Acid PretreatmentY_(Ara/w) 0.005 ± 0.002(77.2) 0.006 ± 0.001(90)   0.007 ± 0.003(98.2)0.007 ± 0.002(100) 0.473 pretreatment liquid Y_(Gal/w) 0.009 ±0.004(95.7) 0.012 ± 0.004(120) 0.011 ± 0.002(117) 0.010 ± 0.004(100)0.645 Y_(Glu/w) 0.043 ± 0.016(80.7)  0.044 ± 0.020(82.2)  0.048 ±0.023(91.1) 0.053 ± 0.014(100) 0.356 Y_(Xyl/w) 0.079 ± 0.036(74.9) 0.085 ± 0.028(80.8)  0.103 ± 0.047(97.2) 0.106 ± 0.030(100) 0.300Y_(Man/w) 0.010 ± 0.005(82)   0.010 ± 0.003(80.8)  0.012 ± 0.006(99.7)0.012 ± 0.003(100) 0.464 Y_(acetic acid) 0.072 ± 0.021(117)  0.080 ±0.024(131) 0.070 ± 0.007(114) 0.061 ± 0.012(100) 0.175 Enzyme Y_(Ara/w)0.0004 ± 0.00(179)  0.0003 ± 0.00(113) 0.0003 ± 0.00(123) 0.0002 ±0.00(100) 0.443 hydrolysate Y_(Gal/w)  0.001 ± 0.00(89.6)  0.001 ±0.00(93.6)  0.001 ± 0.00(88.8)  0.001 ± 0.00(100) 0.395 Y_(Glu/w)  0.318± 0.03(106)  0.302 ± 0.04(101)  0.314 ± 0.03(105)  0.298 ± 0.02(100)0.477 Y_(Xyl/w) 0.005 ± 0.001(125)  0.006 ± 0.002(154) 0.005 ±0.002(133) 0.004 ± 0.001(100) 0.151 Y_(Man/w) 0.003 ± 0.001(90.1) 0.003± 0.001(102) 0.003 ± 0.004(106)  0.003 ± 0.00(100) 0.916 PretreatmentY_(Ara/w) 0.005 ± 0.002(80.8) 0.006 ± 0.001(91)   0.007 ± 0.003(99.1)0.007 ± 0.002(100) 0.515 liquid + Y_(Gal/w) 0.010 ± 0.004(94.7) 0.013 ±0.004(116) 0.012 ± 0.002(113) 0.011 ± 0.003(100) 0.695 Enzyme Y_(Glu/w)0.361 ± 0.029(102)   0.346 ± 0.049(98.4) 0.363 ± 0.018(103) 0.351 ±0.020(100) 0.798 hydrolysate Y_(Xyl/w) 0.084 ± 0.036(76.8)  0.091 ±0.028(83.6)  0.108 ± 0.046(98.6) 0.109 ± 0.030(100) 0.344 Y_(Man/w)0.013 ± 0.006(84)  0.013 ± 0.003(86)  0.016 ± 0.005(101) 0.015 ±0.003(100) 0.468 Pretreatment Y_(Hexoses/W) 0.385 ± 0.029(101)   0.373 ±0.053(98.4) 0.392 ± 0.015(103) 0.378 ± 0.023(100) 0.850 liquid +Y_(Pentoses/W) 0.090 ± 0.039(77.0)  0.098 ± 0.030(84.0)  0.115 ±0.050(98.6) 0.116 ± 0.032(100) 0.352 Enzyme Y_(Monosaccharides/W) 0.475± 0.054(95.6)  0.471 ± 0.070(95.0) 0.507 ± 0.055(102) 0.495 ± 0.049(100)0.630 hydrolysate

Carbohydrate Content Analysis:

FIG. 2 shows the carbohydrate contents of transgenic and wild-typehybrid aspens. The differences between the transgenic lines and thewild-type were small. This agrees with Py-GC-MS data, which also showlittle difference between transgenic lines and wild-type (Table 16). Inconclusion, as the differences in chemical composition were small, thecellulose of the transgenic lines is significantly more susceptible toenzymatic cleavage and deconstruction of the polymeric wood structure.

TABLE 16 Py-GC/MS analysis of transgenic lines and wild-type. Cell wallComposition (%) #H-H12.1 [5] #H-H12.2 [5] #H-H12.3 [3] T89 Carbohydrate83.72 ± 1.57(101)  82.72 ± 1.28(99.9)  83.35 ± 1.88 (100)  82.74 ± 1.57(100)  related Lignin 15.21 ± 1.55(93.5)  16.22 ± 1.26(99.8)  15.63 ±1.86(96.1)  16.25 ± 1.56(100)  S 8.31 ± 0.96(93.0) 9.44 ± 0.70(105) 8.54 ± 1.33(95.6) 8.93 ± 1.12(100) G 5.34 ± 0.60(95.0) 5.35 ± 0.42(95.1)5.30 ± 0.30(94.1) 5.62 ± 0.51(100) H 1.30 ± 0.24(91.4) 1.17 ± 0.24(82.0)1.53 ± 0.26(107)  1.43 ± 0.30(100) S/G 1.55 ± 0.11(97.8) 1.76 ±0.045(110)*  1.60 ± 0.17 (101)  1.60 ± 0.16(100) P 0.24 ± 0.01(92.2)0.26 ± 0.04(98.9) 0.25 ± 0.03(97.2) 0.26 ± 0.04(100)

Example 12: Growth of Arabidopsis thaliana is Enhanced by Expression ofa Transgene Encoding an STT632 Ortholog

Based on phylogenetic analysis, the ortholog to STT632 (TF0137)corresponds to the gene AT2G38470 in Arabidopsis thaliana with thenucleic acid sequence SEQ ID No: 53. This ortholog gene, encoding theamino acid sequence SEQ ID No: 54, was cloned under the control of the35S promoter creating the construct AtTF0137, which was over-expressedin plants.

Methods

Cloning the AT2G38470 Gene:

Based on its known sequence, the coding sequence of the Arabidopsisthaliana AT2G38470 gene was synthesized (Genscript), flanked byrecombination sites for subsequent Gateway cloning. The synthesized genewas subcloned into the binary over-expression vector pK2GW7 usingGateway LR recombination cloning (Invitrogen), where the gene was placedunder the control of the CaMV 35S promoter. The cloned gene was verifiedusing restriction digestion of the final pK2GW7 vector with insert andby sequencing.

Plant Transformation:

The construct, AtTF0137, were transformed into Arabidopsis thalianacol-0 with the transformation method Floral dip.

Plant Growth:

The transgenic Arabidopsis thaliana lines of AtTF0137, were growntogether with their wild-type control (col-0) plants, in a growthchamber, short days (8 h). The plants were fertilized weekly. The plantswere grown for 3 weeks before harvest. During this time the diameter ofthe rosettes was measured once a week.

Results

The measured diameter of the rosettes of the transgenic Arabidopsisthaliana plants transformed with the selected gene is presented in thetable below. The two lines, AtTF0137-line 2 and -line 4, showedsignificantly increased growth as compared to wt col-0 plants.

TABLE 17 Increased growth of Arabidopsis thaliana. Rosette diameterafter 3 weeks Line against col-1 Line name Average Stedv t-testAtTF0137- line 1 5.3 1.1 0.14 AtTF0137- line 2 7.9 1.6 0.05 AtTF0137-line 3 4.4 0.9 0.03 AtTF0137- line 4 8.6 1.0 0.01 AtTF0137- line 5 5.20.5 0.01 Col-1 6.4 1.1

ADDITIONAL ASPECTS

Aspect A1. A method for producing a genetically modified woody planthaving increased biomass and/or wood density compared to a correspondingnon-genetically modified woody plant of the same species, said methodcomprising:a) enhancing the level of expression of at least one polypeptide havingan amino acid sequence selected from among SEQ ID NO.: 2, 28 and 38 oran ortholog thereof in a woody plant, a woody plant cell or a partthereof, and/orreducing the expression of at least one polypeptide having an amino acidsequence selected from among SEQ ID NO.: 58, 74, 88, 98, 106 and 128 oran ortholog or paralog thereof in a woody plant, a woody plant cell or apart thereof;b) generating and/or selecting a woody plant, woody plant cell or a partthereof with improved biomass and/or wood density as compared to acorresponding non-genetically modified woody plant; andc) growing the woody plant, the woody plant cell or the part thereofunder conditions which permit development of a woody plant.Aspect A2. The method according to aspect A1, further comprising:d) selfing or crossing the genetically modified woody plant with itselfor another woody plant to produce seed; ande) growing a progeny woody plant from the seed, wherein the progenywoody plant has increased biomass and/or wood density.Aspect A3. The method according to aspect A1 or A2 wherein the at leastone polypeptide is selected from the group consisting of:a) a polypeptide having an amino acid sequence selected from among SEQID NO.: 2, 28, 38, 58, 74, 88, 98, 106 and 128;b) an ortholog polypeptide to the polypeptide having SEQ ID NO: 2,wherein the amino acid sequence of said ortholog polypeptide has atleast 70% sequence identity to a sequence selected from among SEQ ID NO:2, 4, 6, 8, 10, 12, 14, 16, 18, 20, 22, 24 and 26;c) an ortholog polypeptide to the polypeptide having SEQ ID NO: 28,wherein the amino acid sequence of said ortholog polypeptide has atleast 70% sequence identity to a sequence selected from among SEQ ID NO:28, 30, 32, 34 and 36;d) an ortholog polypeptide to the polypeptide having SEQ ID NO: 58,wherein the amino acid sequence of said ortholog polypeptide has atleast 70% sequence identity to a sequence selected from among SEQ ID NO:58, 60, 62, 64, 66, 68, 70 and 72;e) an ortholog polypeptide to the polypeptide having SEQ ID NO: 74,wherein the amino acid sequence of said ortholog polypeptide has atleast 70% sequence identity to a sequence selected from among SEQ ID NO:74, 76, 78, 80, 82, 84 and 86;f) an ortholog polypeptide to the polypeptide having SEQ ID NO: 88,wherein the amino acid sequence of said ortholog polypeptide has atleast 70% sequence identity to a sequence selected from among SEQ ID NO:88, 90, 92, 94 and 96;g) an ortholog polypeptide to the polypeptide having SEQ ID NO: 98,wherein the amino acid sequence of said ortholog polypeptide has atleast 70% sequence identity to a sequence selected from among SEQ ID NO:98, 100, 102 and 104;h) an ortholog polypeptide to the polypeptide having SEQ ID NO: 106,wherein the amino acid sequence of said ortholog polypeptide has atleast 70% sequence identity to a sequence selected from among SEQ ID NO:106, 108, 110, 112, 114, 116, 118, 120, 122, 124 and 126; andi) an ortholog polypeptide to the polypeptide having SEQ ID NO: 128,wherein the amino acid sequence of said ortholog polypeptide has atleast 70% sequence identity to a sequence selected from among SEQ ID NO:128, 130, 132, 134, 136 and 138.Aspect A4. The method according to any one of aspects A1 to A3 whereinthe polypeptide is encoded by any one of:a) a nucleic acid molecule having a nucleotide sequence selected fromamong SEQ ID NO: 1, 27, 37, 57, 73, 87, 97, 105 and 127;b) a nucleic acid molecule having a nucleotide sequence selected fromamong SEQ ID NO: 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29,31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65,67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, 91, 93, 95, 97, 99, 101,103, 105, 107, 109, 110, 113, 115, 117, 119, 121, 123, 125, 127, 129,131, 133, 135, and 137; andc) a polynucleic acid which hybridizes under stringent hybridizationconditions to any one nucleic acid molecule of (a) or (b).Aspect A5. The method according to any one of aspects A1 to A3, whereinthe step of reducing the expression of the at least one polypeptidecomprises at least one of:a) introducing into at least one woody plant cell a nucleic acidmolecule encoding a ribonucleic acid sequence, which is able to form adouble-stranded ribonucleic acid molecule, whereby a fragment of atleast 17 nucleotides of said double-stranded ribonucleic acid moleculehas a nucleic acid sequence having at least 70% nucleic acid sequenceidentity to any one of SEQ ID NO: 57, 73, 87, 97, 105 and 127;b) introducing into at least one woody plant cell an RNAi or antisensenucleic acid molecule, whereby the RNAi or antisense nucleic acidmolecule comprises a fragment of at least 17 nucleotides with a nucleicacid sequence having at least 70% nucleic acid sequence identity to anyone of SEQ ID NO: 57, 73, 87, 97, 105 and 127; andc) introducing into at least one woody plant cell a nucleic acidconstruct able to recombine with and silence, inactivate, or reduce theexpression of an endogenous gene, wherein the gene comprises anucleotide sequence selected from among SEQ ID NO: 57, 73, 87, 97, 105and 127;d) introducing or inducing a non-silent mutation in an endogenous geneto silence, inactivate, or reduce expression of the gene, wherein thegene comprises a nucleotide sequence selected from among SEQ ID NO: 57,73, 87, 97, 105 and 127; ande) T-DNA inactivation of at last one endogenous gene, wherein the genecomprises a nucleotide sequence selected from among SEQ ID NO: 57, 73,87, 97, 105 and 127.Aspect A6. The method according to any one of aspects A1 to A3, whereinthe step of enhancing the expression of at least one polypeptidecomprises introducing into at least one woody plant cell:a) at least one nucleic acid molecule encoding a polypeptide, whereinthe amino acid sequence of the polypeptide is selected from among SEQ IDNO: 2, 28 and 38; orb) at least one nucleic acid molecule, wherein the nucleotide sequenceof the molecule is selected from among SEQ ID NO: 1, 27 and 37; andc) at least one regulatory sequence operably linked to the at least onenucleic acid molecule of (a) or (b).Aspect A7. The method according to aspect A6 further comprising:d) providing a vector comprising the at least one nucleic acid molecule(a) or (b), and at least one regulatory sequence (c); ande) transforming at least one woody plant cell with the vector.Aspect A8. The method according to any one of aspect A1 to A7, whereinthe genetically modified woody plant is a hardwood tree selected fromthe group consisting of acacia, eucalyptus, hornbeam, beech, mahogany,walnut, oak, ash, willow, hickory, birch, chestnut, poplar, alder,aspen, maple, sycamore, ginkgo, a palm tree, sweet gum, cypress, Douglasfir, fir, sequoia, hemlock, cedar, juniper, larch, pine, redwood, spruceand yew.Aspect B1. A genetically modified woody plant having increased biomassand/or wood density as compared to a corresponding non-geneticallymodified woody plant of the same species, wherein said geneticallymodified woody plant expresses enhanced levels of at least onepolypeptide having an amino acid sequence selected from among SEQ IDNO.: 2, 28 and 38 or an ortholog thereof, and/orexpresses reduced levels of at least one polypeptide having an aminoacid sequence selected from among SEQ ID NO.: 58, 60, 62, 64, 66, 68, 70and 72, or an ortholog thereof.Aspect B2. The genetically modified woody plant according to aspect B1,wherein the genome of said woody plant comprises a genetic modificationselected from any one of:a) a non-silent mutation in at least one endogenous gene having anucleotide sequence of any one of SEQ ID No: 57, 73, 87, 97, 105 and 127that silences or reduces expression of the gene; orb) a transgene inserted into said genome, said transgene comprising anucleic acid molecule encoding a ribonucleic acid sequence, which isable to form a double-stranded ribonucleic acid molecule, whereby afragment of at least 17 nucleotides of said double-stranded ribonucleicacid molecule has a nucleic acid sequence having at least 70% sequenceidentity to a nucleic acid molecule of any one of SEQ ID No: 139-144; orc) a transgene inserted into said genome, said transgene comprising atleast one nucleic acid molecule having a nucleotide sequence selectedfrom among SEQ ID No: 1, 27, and 37 and at least one regulatory nucleicsequence fused to and controlling expression of said at least onenucleic acid molecule.Aspect B3. The genetically modified woody plant according to aspect B1,wherein said woody plant has an increased expression of at least one ofpolypeptide, wherein the amino acid sequence of said polypeptide has atleast 70% amino acid sequence identity to a sequence selected among SEQID No.: 2, 28 and 38.Aspect B4. The genetically modified woody plant according to aspect B1,wherein said woody plant has a reduced expression of at least one ofsaid polypeptides, wherein the amino acid sequence of said polypeptidehas at least 70% sequence identity to a sequence selected among SEQ IDNO.: 58, 60, 62, 64, 66, 68, 70, 72, 74, 76, 78, 80, 82, 84, 86, 88, 90,92, 94, 96, 98, 100, 102, 104, 106, 108, 1110, 112, 114, 116, 118, 120,124, 126, 128, 130, 132, 134, 136 and 138.Aspect B5. The genetically modified woody plant according to any one ofaspects B1-B4, wherein the genetically modified woody plant is selectedfrom the group consisting of acacia, eucalyptus, hornbeam, beech,mahogany, walnut, oak, ash, willow, hickory, birch, chestnut, poplar,alder, aspen, maple, sycamore, ginkgo, a palm tree, sweet gum, cypress,Douglas fir, fir, sequoia, hemlock, cedar, juniper, larch, pine,redwood, spruce and yew.

1. (canceled)
 2. A method for producing a genetically modified woodyplant having increased biomass and/or wood density compared to acorresponding non-genetically modified woody plant of the same species,said method comprising: a) enhancing the level of expression of at leastone polypeptide having an amino acid sequence selected from among SEQ IDNO.: 28 and 38 or an ortholog thereof in a woody plant, a woody plantcell or a part thereof, and/or reducing the expression of at least onepolypeptide having an amino acid sequence selected from among SEQ IDNO.: 58, 74, 88, 98, 106 and 128 or an ortholog or paralog thereof in awoody plant, a woody plant cell or a part thereof; b) generating and/orselecting a woody plant, woody plant cell or a part thereof withimproved biomass and/or wood density as compared to a correspondingnon-genetically modified woody plant; and c) growing the woody plant,the woody plant cell or the part thereof under conditions which permitdevelopment of a woody plant.
 3. The method according to claim 2,further comprising: d) selfing or crossing the genetically modifiedwoody plant with itself or another woody plant to produce seed; and e)growing a progeny woody plant from the seed, wherein the progeny woodyplant has increased biomass and/or wood density.
 4. The method accordingto claim 2, wherein the at least one polypeptide is selected from thegroup consisting of: a) a polypeptide having an amino acid sequenceselected from among SEQ ID NO.: 28, 38, 58, 74, 88, 98, 106 and 128; b)an ortholog polypeptide to the polypeptide having SEQ ID NO: 28, whereinthe amino acid sequence of said ortholog polypeptide has at least 70%sequence identity to a sequence selected from among SEQ ID NO: 28, 30,32, 34 and 36; c) an ortholog polypeptide to the polypeptide having SEQID NO: 38, wherein the amino acid sequence of said ortholog polypeptidehaving at least 70% amino acid sequence identity to a sequence selectedfrom among SEQ ID NO: 38, 40, 42, 44, 46, 48, 52, 54 and 56; d) anortholog polypeptide to the polypeptide having SEQ ID NO: 58, whereinthe amino acid sequence of said ortholog polypeptide has at least 70%sequence identity to a sequence selected from among SEQ ID NO: 58, 60,62, 64, 66, 68, 70 and 72; e) an ortholog polypeptide to the polypeptidehaving SEQ ID NO: 74, wherein the amino acid sequence of said orthologpolypeptide has at least 70% sequence identity to a sequence selectedfrom among SEQ ID NO: 74, 76, 78, 80, 82, 84 and 86; f) an orthologpolypeptide to the polypeptide having SEQ ID NO: 88, wherein the aminoacid sequence of said ortholog polypeptide has at least 70% sequenceidentity to a sequence selected from among SEQ ID NO: 88, 90, 92, 94 and96; g) an ortholog polypeptide to the polypeptide having SEQ ID NO: 98,wherein the amino acid sequence of said ortholog polypeptide has atleast 70% sequence identity to a sequence selected from among SEQ ID NO:98, 100, 102 and 104; h) an ortholog polypeptide to the polypeptidehaving SEQ ID NO: 106, wherein the amino acid sequence of said orthologpolypeptide has at least 70% sequence identity to a sequence selectedfrom among SEQ ID NO: 106, 108, 110, 112, 114, 116, 118, 120, 122, 124and 126; and i) an ortholog polypeptide to the polypeptide having SEQ IDNO: 128, wherein the amino acid sequence of said ortholog polypeptidehas at least 70% sequence identity to a sequence selected from among SEQID NO: 128, 130, 132, 134, 136 and
 138. 5. The method according to claim2, wherein the polypeptide is encoded by any one of: a) a nucleic acidmolecule having a nucleotide sequence selected from among SEQ ID NO: 27,37, 57, 73, 87, 97, 105 and 127; b) a nucleic acid molecule having anucleotide sequence selected from among SEQ ID NO: 27, 29, 31, 33, 35,37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71,73, 75, 77, 79, 81, 83, 85, 87, 89, 91, 93, 95, 97, 99, 101, 103, 105,107, 109, 110, 113, 115, 117, 119, 121, 123, 125, 127, 129, 131, 133,135, and 137; and c) a polynucleic acid which hybridizes under stringenthybridization conditions to any one nucleic acid molecule of (a) or (b).6. The method according claim 2, wherein the step of reducing theexpression of the at least one polypeptide comprises at least one of: a)introducing into at least one woody plant cell a nucleic acid moleculeencoding a ribonucleic acid sequence, which is able to form adouble-stranded ribonucleic acid molecule, whereby a fragment of atleast 17 nucleotides of said double-stranded ribonucleic acid moleculehas a nucleic acid sequence having at least 70% nucleic acid sequenceidentity to any one of SEQ ID NO: 57, 73, 87, 97, 105 and 127; b)introducing into at least one woody plant cell an RNAi or antisensenucleic acid molecule, whereby the RNAi or antisense nucleic acidmolecule comprises a fragment of at least 17 nucleotides with a nucleicacid sequence having at least 70% nucleic acid sequence identity to anyone of SEQ ID NO: 57, 73, 87, 97, 105 and 127; and c) introducing intoat least one woody plant cell a nucleic acid construct able to recombinewith and silence, inactivate, or reduce the expression of an endogenousgene, wherein the gene comprises a nucleotide sequence selected fromamong SEQ ID NO: 57, 73, 87, 97, 105 and 127; d) introducing or inducinga non-silent mutation in an endogenous gene to silence, inactivate, orreduce expression of the gene, wherein the gene comprises a nucleotidesequence selected from among SEQ ID NO: 57, 73, 87, 97, 105 and 127; ande) T-DNA inactivation of at last one endogenous gene, wherein the genecomprises a nucleotide sequence selected from among SEQ ID NO: 57, 73,87, 97, 105 and
 127. 7. The method according to claim 2, wherein thestep of enhancing the expression of at least one polypeptide comprisesintroducing into at least one woody plant cell: a) at least one nucleicacid molecule encoding a polypeptide, wherein the amino acid sequence ofthe polypeptide is selected from among SEQ ID NO: 28 and 38; or b) atleast one nucleic acid molecule, wherein the nucleotide sequence of themolecule is selected from among SEQ ID NO: 27 and 37; and c) at leastone regulatory sequence operably linked to the at least one nucleic acidmolecule of (a) or (b).
 8. The method according to claim 7, furthercomprising: d) providing a vector comprising the at least one nucleicacid molecule (a) or (b), and at least one regulatory sequence (c); ande) transforming at least one woody plant cell with the vector.
 9. Themethod according to claim 2, wherein the genetically modified woodyplant is a hardwood tree selected from the group consisting of acacia,eucalyptus, hornbeam, beech, mahogany, walnut, oak, ash, willow,hickory, birch, chestnut, poplar, alder, aspen, maple, sycamore, ginkgo,a palm tree, sweet gum, cypress, Douglas fir, fir, sequoia, hemlock,cedar, juniper, larch, pine, redwood, spruce and yew.
 10. A geneticallymodified woody plant having increased biomass and/or wood density ascompared to a corresponding non-genetically modified woody plant of thesame species, wherein said genetically modified woody plant expressesenhanced levels of at least one polypeptide having an amino acidsequence selected from among SEQ ID NO.: 28 and 38 or an orthologthereof, and/or expresses reduced levels of at least one polypeptidehaving an amino acid sequence selected from among SEQ ID NO.: 58, 60,62, 64, 66, 68, 70 and 72, or an ortholog thereof.
 11. The geneticallymodified woody plant according to claim 10, wherein the genome of saidwoody plant comprises a genetic modification selected from any one of:a) a non-silent mutation in at least one endogenous gene having anucleotide sequence of any one of SEQ ID No: 57, 73, 87, 97, 105 and 127that silences or reduces expression of the gene; or b) a transgeneinserted into said genome, said transgene comprising a nucleic acidmolecule encoding a ribonucleic acid sequence, which is able to form adouble-stranded ribonucleic acid molecule, whereby a fragment of atleast 17 nucleotides of said double-stranded ribonucleic acid moleculehas a nucleic acid sequence having at least 70% sequence identity to anucleic acid molecule of any one of SEQ ID No: 139-144; or c) atransgene inserted into said genome, said transgene comprising at leastone nucleic acid molecule having a nucleotide sequence selected fromamong SEQ ID No: 1, 27, and 37 and at least one regulatory nucleicsequence fused to and controlling expression of said at least onenucleic acid molecule.
 12. The genetically modified woody plantaccording to claim 10, wherein said woody plant has an increasedexpression of at least one of polypeptide, wherein the amino acidsequence of said polypeptide has at least 70% amino acid sequenceidentity to a sequence selected among SEQ ID No.: 28 and
 38. 13. Thegenetically modified woody plant according to claim 10, wherein saidwoody plant has a reduced expression of at least one of saidpolypeptides, wherein the amino acid sequence of said polypeptide has atleast 70% sequence identity to a sequence selected among SEQ ID NO.: 58,60, 62, 64, 66, 68, 70, 72, 74, 76, 78, 80, 82, 84, 86, 88, 90, 92, 94,96, 98, 100, 102, 104, 106, 108, 1110, 112, 114, 116, 118, 120, 124,126, 128, 130, 132, 134, 136 and
 138. 14. The genetically modified woodyplant according to claim 10, wherein the genetically modified woodyplant is selected from the group consisting of acacia, eucalyptus,hornbeam, beech, mahogany, walnut, oak, ash, willow, hickory, birch,chestnut, poplar, alder, aspen, maple, sycamore, ginkgo, a palm tree,sweet gum, cypress, Douglas fir, fir, sequoia, hemlock, cedar, juniper,larch, pine, redwood, spruce and yew.